CA2236295A1 - Ultralow background multiple photon detector - Google Patents

Abstract

An ultralow background multiphoton detector device for radioisotopes (20) has a background of about one count per day and can detect one attomole of material. Two opposed gamma and X-ray photon detectors each preferably include a scintillation crystal (22) and a photomultiplier tube (26). Sandwiched between the detectors are a separator (32) and a sample holder (34) for a sample (36) labelled with a radioisotope such as I125. Each detector converts emissions from the radioisotope into electric output pulses in a base (30) which pulses are then subjected to signal processing for pulse shape analysis, discrimination of coincident from non-coincident events, and quantification of the radioisotope. Detector materials and components are selected to minimize background, and are shielded from external radiation in a composite shield (55). Detectors operating on different principes and sample holders of different configurations may be used. The device may further be constructed to resolve and provide images of two-dimensional sample arrays.

Description

CA 0223629~ 1998-04-29 ULTRALOW BACKGROUND MULTIPLE PHOTON DETECTOR

BACKGROUND OF THE INVENTION
Field This invention relates to multiphoton radioisotope detectors with ultralow background.
S These detectors can ~luallliry coincident gamma and X-ray emissions from electron capture (EC) isotopes, combining coincident counting and other background rejection measures to achieve extraoldil~ly sensitivity.
Back,eround Il,rollllalioll Hal.lwal~ impl~m~nt~l coincidence counting is used in a detector for positron-gamma (pg) ellliLIel~, disclosed in United States Patent No. 5,083,026. Within 10 nanoseconds after the coinri~lent emission of a ~O~,iL.oll and a g~mm~ the positron ~nnihil~t~ the electron producing two back-to-back gamma photons with energies of Sll keV. Multiple scintillation detectors are used to register the three coincident high energy (E 2 250 keV) g~mm~c, and events lacking this triple gamma cign~tllre are rejected. These i~sLlu~llerl~, have serious 15 limit~til~ns, in particular the type of isotopes that may be used and the large mass and high cost of the scintillator crystals.
According to WO 9S/10308, corresponding to U.S. Patent No. 5,532,122, the disclosure of which is hereby incorporated by reference, a method is disclosed for coincident l ion of gamma and X-ray emitting isotopes at low qn~ntitiPs with ultralow background 20 on the order of one count per hour.
Simlllt~n~ous counting of individual and coincident gamma and x-rays is taught in Oe~,le~ et al., U.S. patent 4,005,292; Horrocks et al., U.S. patent 4,016,418; Coffey, U.S.
patent 3,974,088; and Fymat et al., U.S. Patent 4,682,604. However, these are all limited to high radioactivity applications.

It is an object of the invention to provide an ultralow background detector which is specially targeted for low levels of coincident gamma/X-ray emitting isotopes (CGX isotopes).
It is a further object to use the coincident photons origin~ting from distinct nuclear and electron shell excitations of CGX isotopes as a means to selectively quantify CGX events and 30 achieve background rejection.
A further object of the invention is to reduce background to less than one count per hour by eli~inA~ g all .si~ni~lr~nt sources of background. It is a further objective of the CA 0223629~ 1998-04-29 invention to overcome the hlh~ lly lower detection efficiency of the selective CGX counting mode as compared to single gamma counl~ls, to provide a sensilivily to sub-pico Curie samples, and an ability to detect less than 5 x 10-2l mole of labeled molecules, a zeptomole/ml, or even a single large labelled biomolecule, with detection efficiency greater than 10% and 5 reproducibility of about 1%.
It is another object to provide an instrument that can operate either in a non-coincident (single photon) mode or a coincident gamma-x-ray emissions (CGX events) counting mode, with enormous dynamic range and linearity of better than 5 % over nine orders of m~gnhlllle.
Detectors according to the invention satisfy these objects. They have multicolor ability 10 in that several isotopes can be measured and ~ ;"g,li~h~cl in the same sample. They can easuie many St;~alf; samples and yield very reproducible results. They incorporate sellf-calibration and self-diagnostics. They can provide spatial resolution of 100 micrometers or lower. All these advantages are accomplished in transportable, ill~ llsive devices much smaller than conventional devices. Rec~-lse of the h~lcased sel~iliviLy of ii~iLlulllellL~
15 according to the invention, isotope concellLlalions may be reduced, thus lowering isotope acquisition costs, exposure of personnel, and radioactive waste disposal problems.
A Multi-Photon Detection a~ LuS (MPD) for detecting a radioisotope (cohlcidelll gamma/X-ray (CGX) emitter) in a sample comprises means for detecting coincident (e.g., gamma and X-ray) radiation from the CGX emitter as output pulses in sepala~e radiation 20 ~etectors, means for analyzing the shape and height of the pulses on-line to identify pulses char~cteri~tic of single photon emissions, means for discrimin~ting and rejecting (e.g., non-coincident gamma and X-ray radiation) spurious pulses, means for ~u~l.lcssillg background radiation, preferably a composite radiation shield and a sepdl~or that absorbs X-rays, and means for quallliryiilg the pl_sel1ce of the (e.g., CGX) emitter in the sample in an amount of 25 less than about 100 picoCurie.
Further objectives and advantages will become a~a cllL from a consideration of the detailed description and drawings.

BRIEF DESCRIPTION OF THE FIGURES
The invention is better understood by reading the following det~ d description with 30 le~rellce to the accolllp~lyillg figures, in which:
- Figure 1 shows a block diagram of multiphoton detector.
Figure 2 shows a detector sl-ba~embly.

CA 0223629~ 1998-04-29 Figure 3 is a block diagram of an integrated r~hot-mllltirlier base including amplifier and high voltage power supply. Figures 3A to 3C show exemplary embo~l;,-,P~ of the component parts in more detail. Figure 3A shows a PMT base; Figure 3B shows a negative high voltage power supply; and Figure 3C shows an amplifier and shaper.
Figure 4 graphs the dynamic range of an MPD according to the invention.
Figure S shows the dynamic range of an MPD device compared to a color specl,u,lleter.
Figure 6 shows the layout for a se~lPntiAl sample MPD with a sample changer.
Figure 7 is a block diagram of an MPD embodiment acording to the invention.
Figures 8a, 8b and 8c are a flow chart of a method according to an embodiment of the invention.
DETAILED DESCRIPTION OF THE INVENTION
A typical embodiment of an MPD device system according to the invention is shownin Figure 1. This exemplary system as illustrated has five major cc",~o~e"L~, including:
1. two photon detectors, e.g., scintillator/photosensor modules;

2. ~hilo!rling co,l,po~ including a ~e~alaLor, 3. a sample holder sandwiched between the photon dele~;lol~;

4. detection ele~;Llol~ics including voltage supplies; and 5. signal proce~sing for data acquisition, pulse shape/height analysis, and display.
20 All of the components of the exeTnrlAry embodiment are selectçd and alldnged to ,in;--,i~.o radioactive background.
In detector sub-assembly 20, two scintillation crystals 22 with applo~uliate photosensors 26, e.g., PMT's, face each other on s~lbst~ntiAlly a common axis. They are separated by an app,upliat~ se~aldlor 32, preferably less than 8 mm thick with a centered biconical hole 34 allowing sample 36 held therein to irradiate both crystals 22. The dimensions of hole 34 are chosen so that the solid angle of the opening just encomp~ses the faces of crystals 22.
Stllaldlol 32 optimally limits cross-talk beLweell crystals 22 and the thickness is selected to limit certain background events. The detectors are placed in composite shield 55 to ~limini~h background due to ambient radioactivity. This shield, depending on applications, can be from 0.5 to 20 kg in weight. Each photosensor 26 is also provided with ~hie!.1ing 27.In operation, energy deposited within scintillation crystals 22 is converted into charge pulses by photosensors 26 and then into voltage pulses in integrated base 30. The pulses are shaped by fast low-noise preamplifiers 52, as shown in detail in Figures 3A-3C. The base CA 0223629~ 1998-04-29 electronics also include high-voltage power supply 50 for the photosensors 26, and high-gain amplifiers 54. Special "Lliall~ular shape" shaping preamplifiers 52 are used to permit both fast coincidence and good energy resolution. The signals A and B from both detectors 26, as well as the coincidence trigger A+B, are inputs to a PC-based digital storage oscilloscope (DSO) card 58, which is used for fast real-time pulse-shape/pulse-height analysis under software control.
The software supporting the DSO card-based pulse-shape analysis on the PC is preferably coded in Borland Turbo Pascal and C++, which helps make the code highly modular, llans~eLll, and easy to ~;u~lOllli~. Through changes in the software, the same device can be adapted for a variety of applications willwuL har-l~alc; changes. The most time-critical procedures are preferably coded in IBM PC assembly language to maximize effiriPnry of acquiring and analyzing signals.
DSO card 58 has two inputs with at least 8-bit analog-to-digital conv~ (ADC's) and an additional input, which is used as an acquisition trigger. It is able to monitor the two input channels siml~lt~n~ously at a sampling rate of at least 10 MHz. The signals are acquired at a sampling rate of up to 100 mPg~c~mples/sec and are continuously stored in on-board memory until a trigger is ~letectPA, wh~l~u~oll a pre-defined llulllber of post-trigger data points are acquired. Once the acquisition is stopped, the contents of the on-board memory become available for the host colll~ul~l CPU 59.
Any of the DSO ch~nnPls can be used as the trigger, or triggering can be done through software and/or trigger ele~;llol~ics 56. The sampling rate, the number of post-trigger points, the gains on the channels, and the triggering conditions can be set through a PC I/0 port, while the on-board memory can be ~ccessecl directly through a 4 Kbyte window, which allows fast retrieval using the CPU's string instructions or DMA ~ r~r to other peripheral devices, for example.
The pulse shape analysis functions as a way to discriminate pulses due to radioactive decay from spurious pulses based on the following charactPri~tir~. When a photon is absorbed by a scintillator/PMT or scintillator/photodiode combination, it produces a pulse with a characteristic shape and a predictable amplitude proportional to the energy deposited by the photon in the scintillator. In contrast, the shapes of spurious pulses electrom~nt-tic~lly in~1uce-1 in the readout electronics are not so well defined. Spurious pulses include single - narrow spikes, rapid successions of such spikes, random electronic noise and PMT dark pulses. In a pulse-height analyzing system spurious pulses may considerably cont~min~te the CA 0223629~ 1998-04-29 wo 97/16746 PCT/US96/16968 acquired spectra and dimini~h the system's overall signal-to-backgl~,ulld ratio. Also, many "real" pulses caused by detection of photons could be di~ol~d, e.g., when two sepala~ events occur within a short interval of time (pile-up artifacts) or a photon-caused pulse is distorted by a coincident elecL~ gn~ti~ pick-up. Such events blur the acquired s~,ectlu ll, and are S either discarded or correctly illte~ led based on shape analysis. High-frequency electronic noise in the front end electronics pelLull,s the spec~ , decreasing energy resolution.
However, through pulse-shape analysis the spectrum can be enh~nred to the actual resolution of the photosensor.
Design considerations for reducing background will next be ~ c~lsse(l The following 10 sources of back~lou-ld are signifi~antly reduced in the exemplary embo~lim~nt~ according to the invention:
Al radioactive co--~ on of scintillator crystals;
A2 radioactive cont~min~tion of ph~ tom~lltiplier tubes (PMT's);
A3 radioactive co"~ tion of shields;
15 A4 high energy g~mm~ from the e~ ilol~el.l;
Bl neutron inflllrecl g~mm~c from the scintill~tors, PMT's and shields;
B2 cosmic ray in~ ced g~mm~ from the scintillators, PMT's and shields;
B3 direct hits due to cosmic rays;
Cl dark pulses of PMT's;
20 C2 cosmic ray in~ ced dark ~;UllcllLS of PMT's;
Dl electronic pick-up;
D2 vibrational pick-up.
The first group of background sources (Al-A4) produces background levels on the order of a few counts per second (cps). Conventional methods of ~i...i.~i~hi.~g this class of background involve use of ultrapure materials, often purified to over 99.999%. However, purification methods are too ~Al,~,nsi~e to be used in low cost instruments for biom~lir~l applications. St~ti~tir~l m~tho.1c of background subtraction are limited by st~ti~tir~l artifacts and can be used only if the signal to background ratio (S/B) is large, say > 10.The second group of background sources (Bl-B3) is l~s~onsible for backgrounds at the 0.1 - 1 counts per minute (cpm) level. Active shielding and sophi~tir~t~cl pulse shape analysis can be used to reject t_is background, but the use of active shields tends to be very e~e~ /e.
Furthermore, the use of active shields often leads to complications in the detector geometry and limitations on the space available for samples, thus decreasing the utility of the device.

CA 0223629~ 1998-04-29 wo 97/16746 PCT/US96/16968 The third group of bac~luulld sources (C1-C2) produces a few counts per hour (cph).
These sources are extremely tliffirlllt to reject by haldw~e means. A ~yllc~ ic combination of coincidence, ha~ ~c means and sophictir~te(l pulse shape analysis permits rejection of about 90% of the background in this class. However, this requires heavy use of on-line pulse shape analysis based on a digital storage oscilloscope.
The fourth group of background sources (D1-D2), electronic and vibrational pick-up noise, is very much system and site clepen~l~nt. Conventional NIM based electronics show typical noise of 0.1 cpm per unit, dependillg on the total number of modules used. Even when using well grounded coaxial cables, with 10 tû 20 cables used to connect a typical NIM
10 system, the observed ele-;Lloi~.~n~otic noise is about 1 cpm. Also, the use of external high voltage sources negatively influences the level of pick-up noise.
To reach the level of less than one spurious count per hour, special low noise electronics are used. There is a trade-off between the on-line colll~ulillg ~lenn~n~1c of a system in accordance with the invention n~cess~ry to ~u~ress the sources B1-B3 and C1-C2, and the 15 reslllting increase in the electronic pick-up noise.
Isotope selection is now tliccllcsecl The EC emillcls most ap~lupliate for use with the invention are listed in U.S. patent 5,532,122, lists 1~. EC elllil~ typically have two coincident photons, of which one is always a rather low energy X-ray emitted due to atomic shell leall~1gelllent. Also, some Auger electrons can be present which can be used 20 advantageously to obtain very high spatial resolution. For the plerclled radioemitter Il25, the second photon also has a low energy; the spectrum has three peaks at 27, 31 and 35 keV, respectively. Designing a MPD detector involves trade-offs between detection efficiency (DE), energy resolution (dE) and ~clllpoldl response. Photon energies considerably influenre the choice of detector el~qnnprlt~. In the case of high energy cnlliLLels with at least one gamma 25 photon with energy higher than 100 keV, heavy hlOlgal~iC scintillators may be practical and economical, whereas for detection of EC isotopes a large class of detectors is available including scintillators, semicon(1ucting detectors and gas detectors.
Preferably, the CGX emitter is an isotope of an element that forms covalent bonds, or one that can be chelated to an organic compound. More preferably the isotope is Il23, I'25, I'25, 30 Br75, or Br77, and most preferably is I'25.
The EC isotope I'25, with a half life of 60 days, is particularly plcrellcd. According to results obtained with older h~Ll~llnellldlion and recorded in nuclear data tables, coincident emission of a 35 keV nuclear gamma and one of several possible X-rays in the 27-31 keV

CA 0223629~ 1998-04-29 range occurs in 7% of the Il25 decays. However, with the improved system of the invention, eA~ .Pnt~ revealed in actuality a coincidence in 25-35% of the decays.
The CGX emitter is preferably an isotope of the family of CGX l~nth~nidP isotopes.
This family inrl~1dPs 54 isotopes with i~lentir~l çhPmir~l plopcllies which are distinguishable 5 based on their dirre~ellt decay photon energies. CGX l~nth~ni-le isotopes can be introduced as labels to biomolecules through the use of chelating groups which capture mPt~llic ions. The ch.omi.~try for ~ uctin~ chP!~ting groups to DNA sllblmit.c and their polymers is well known.
Material selection for an ultralow background CGX system for Il25 will now be ~ Yl~se~. The detection of gamma rays from Il25 requires scintillator de~;tol~ with good 10 energy resolution. Generally, the best energy resolution is achieved with a NaI(Tl) crystal/PMT combination. However, use of NaI(Tl) leads to an additional hard X-ray background due to the characteristic iodine absorption edge. This is be,~ericially eli---i--~led by impleTnPntin~ the plef~ d CaF2/PMT combination accoldil~g to the invention.
All m~teri~l~ used in the CGXD system in more than 1 mg quantity are screened for 15 radioactive hll~ulilies. Glass which contains K40 is preferably replaced by quartz or special low background pyrex. Ferric m~t~ri~l.c are excluded because of radioactive cobalt co..~ ion. Follulldtely some types of plastic, e.g., teflon~9 and acrylic, are açcept~hle, as well as ultrapure copper, tin and lead.
Diagnostics: Diagnostic applications of the invention include DNA sequencing, DNA
20 fingell,lilllillg and diverse forms of Culllp~ ive and/or binding assays, e.g., radioi~ lunoassay (RIA). Using pCi levels of Il25 isotope inputs. exposure to toxins and pathogens which may be resident in assay samples or reagents are a far greater safety concern than the minute amount of Il25 used.
This system can be used to qu~ntit~te sub-attomole ( < 10-l8 M) amounts of 25 biomolecules in diagnostic tests such as IAs, imml-no-PCR and DNA probes, and IRMAs. To obtain an ~lopliate signal to background ratio (S/B) in the face of 1 cps background levels, previous IRMAs have required the use of hazardous qll~ntitiPs of radioisotope. Due to the absence of approved disposal sites, radioactive waste must be stored at the sites that ge~ e the waste (i.e., hospitals, universities and industries), creating potential radiation and bio-30 hazards. According to the invention, solid assay resi~ can be discarded subject only tobiohazard regulations, as they are less radioactive than the ellvilv-llllr~ l bacl~l.,ulld. Liquid residu~l~ will contain most of the radioactivity, but at lower activity than those from current biomP~1ic~l procedures, e.g., radioimmlm~.c~ys (RIA) and typically much belûw the CA 0223629~ 1998-04-29 6 PCT~US96/16968 radioactive level of ground water. Thus, immP~ tP~ disposal will probably be allowed in many cases.
Advantages of a "sandwich" detector geometry are now ~ sed. One element of devices according to the invention is "sandwich" geometry, in which a single sample is placed 5 between two independent scintillator detectors, each read-out by a se~al~tc PMT or equivalent.
Sandwich geometry is coul~ i,lluiLi~e because it leads to twice the number of ele~nPnt~, including PMT's, which are a major source of background. Fùl~ ,lmore, passive shi~ ing is considerably more costly and the overall shape of the detector assembly is somewhat ~wkw~d, clearly ~limini~hing user frien~linPs~. Finally, sandwich geometry flimini~h~s 10 detection efficiency as colll~a~d to well geometry.
However, the use of sandwich geometry lcplesellL~ an optimal trade-off among several re4ui,-,lllcllL~:
* the ability to operate both in OR (non-coincident) and AND (coincident) modes;* geometry l,c~ ;.-g an asylllmeLlic detector configuration;
15 * ...i..;...i,i.lion and/or calibration of absorbtion artifacts; and A major source of background counts in scintillator-based gamma coullL~l~ is radioactive co-.l~ ion of the componellL~ of the detector itself. To reduce the actual radioactive component of the background the volume of the scintillator may be op~ Pcl. The low energy gamma and X-rays associated with I'25 decays are effectively stopped in less than 20 1 mm of NaI(Tl) or 1.5 mm of CaF2(Eu) scintillator. However, producing a well detector with such thin walls is impractical, especially given the need to couple a PMT to the scintillator. Tn~te~-l a flat scintillator geometry is plcrellcd even though it has somewhat lower detection efficiency. In practice, the detection efficiency of 2" diameter flat detectors is about 40% for a small (few mm ~ m~ter) source placed in the middle of the detector. The 25 overall detection efficiency of the system is improved and made less dependent on geometry by using two identical flat round detectors with the source placed be~weell them.
Segmentation of the detector has other benefits as well. It provides the ability in half the cases to distinguish 2-photon events from single photon events with double the photon energy. It also provides the ability to discli,llil~e background by using anti-coincidence 30 rejection teçhni-lues. However, each detector module has a separate PMT, which increases the non-radioactive background component due to PMT dark pulses and various electronic - artifacts. Therefore, effective disclimillation of such non-radioactive events by pulse shape analysis in accordance with the invention is preferred.

CA 0223629~ 1998-04-29 WO 97/16746 PCT~US96/16968 The sandwich detector geometry permits better pelro,lllance than either well detector or flat detector geollleLIies. The qu~lh~tive ~lopelLies of scintillator detectors built with these three dirf~l.,nl geolllel,ies are shown in Table 1.

5Detector Detection Energy Background efficiency resolution Well type ~let~ctQrsvery good (250%) fair poor Flat type detectors poor (<40%) good good Sandwich type good (250%) good excellent detçctQrs Reproducibility and reliability are important realulcs for biomP~1ic~l instruments. In many ~letectQrs~ typical sources of unc~ illly are self-absoll,lion and errors due to variations in the sample positioning inside/in front of the detector. These errors are easier to ~-~i"i,--i,~
when the sample is a liquid, and in this case the use of a well geometry has considerable advantages. Ullrollul~lely, use of liquids leads to non trivial sample h~n~iling problems.
15 Modern di~gnostic meth~ -ls often use formats in which the biological samples are ~ rhecl to a solid surface or are products of a separation process which are adsorbed onto a~l~li~l~
filters. For example, electrophoresis products are trapped in the gel or llol~rc~ d to membranes. Deleclo,s with flat geometry are advantageous when samples with a large surface/volume ratio are used, especially when biological samples are distributed 20 inhomogeneously inside of, or ~tt~rh~d upon, the surface of a solid state filter, support, or membrane with a non-negligible thic~nrc~.
Qu~ntit~tion artifacts can be considerably ~i,--ii,iil-Pd when using two essentially i~ntir~l flat tlet~ctors~ each with independent read-out ele-;llol~ics in accordance with the invention. When self-absorption is negligible and the placement of the sample is correct, both 25 detectors give essentially the same count rate. However, even when the count rate is dirrelelll in both detectors, the sandwich geometry permits the use of highly efficient dirr.,~ iation/compensation srhprnl~s.
There are other advantages to using a sandwich geometry in accordance with the invention, especially for efficient background rejection. Many background events are due to 30 electrom~gnPtic pick-up and dark pulses in PMT's. With two well-separated detectors a CA 0223629~ 1998-04-29 W O 97/16746 PCTrUS96/16968 .cignifir~nt part of the elecllu~ nptir pick-up can be ~1etçcted by col~-p~ g the pulses in both detectors. Thus, a pair of detectors with sepalatc but i(lentic~l electronics can be operated in an anti-coincidence mode to reject electronic and vibrational pick-up.
Background due to K40 co,~ ",i~u~iQn within the PMT's is an hlll)o~ component ofS the total background in a single detector configuration. When two flat detectors are used, the energy deposited in one detector is often very dirrclcnl from the energy deposited in the second detector, which permits rejection of the background due to beta clllillcl~ co"l~"~ ting scintillators, PMT's and shields. The use of a~ropliate scpaldtol~ permits further elimin~tion of background due to high energy photons peneLld~ g from outside the device. It is also 10 useful in lejeclillg cosmic rays.
Another important advantage of the sandwich geometry according to the present invention is its versatility. A prcfc.lcd implement~tion of the invention uses two çssenti~lly irlentir~l detectors for the detection of the isotope I'25. The case of Il25 is somewhat an exception, i.e., multiple photons are emitted in coinri(lenre, but their energies are very close, namely E = 27, 31 and 35 keV. Thus, for I'25, two i(lPntir~l thin scintillators can be used.
However, many other important sources emit multiple photons of quite difr.,~lll el~ies;
often one photon is a soft X-ray (E < 50 keV) while the second photon is a nuclear gamma-ray, e.g., E > 100 keV. For example, this is the case for '23I, which has three lines at 27 keV, 31 keV and 150 keV respectively. In this case, a sandwich detector according to an 20 embodiment of the invention having two scintillators with different crystal thir~n~ss would be optimal. Further, photoswitch elemrnt.~ can be used for detection of photons of ~ignifir~ntly dirr~,lcnl energies.
When using scintillator detectors, it is applopliat~ to physically isolate the scintillators as much as possible to reduce inthlced X-ray crosstalk between them. If the volume of the 25 samples is relatively small (a few hundred microliters) this can be achieved by incorporating a 1-5 mm thick sheet of lead or copper into the sample holder. Isolation of the detectors effectively reduces the background in the single-photon Il25 energy region of interest (ROI) twofold. Detector isolation has much less effect on the background in CaF2(Eu)-based systems because these detectors do not produce secondary X-rays in the I'25 ROI. The use of CaF2(Eu) 30 facilitates construction of MPD detectors in accordance with the invention for samples with large volume. NaI(Tl) or CsI(Tl) based detectors show considerable sensitivity to the geometry and rli~mPter of opening in the sep~lor.

CA 0223629~ 1998-04-29 W O 97/16746 PCT~US96/16968 Co~ aliso~ of a device according to the invention with the detectors at MemorialSloan KettPrin~ Cancer Center (MSKCC) and elsewhere were performed using binary r~ tion~
of commercially available radio-iodinated TSH antibodies. The molecular weight of this antibody is about 40,000 daltons. Most of the tests were performed using a calibration sample 5 set. Additional calibration runs used water dilutions of NaIl2s and Il25-dCTP. For radioactive standards, ethi(~ m bromide was io-lin~ted with Il2s to a specific activity of about 0.1 mCi/mL.
It was subsequ~ntly sequentially diluted by factors of four with isopropyl alcohol. The calibration set includes ten samples, covering the range from a few thousand dpm to about 0.1 dpm. Hundred microliter volumes of each dilution were placed in 200 microliter Eppendorf 10 vials made of a thin plastic.
Results of these comparisons confirrn~d that the MPD permits llRa~ule.lle.lL of lower amounts of radioactivity than other gamma detectors. The detection efficiency and energy resolution of two dirrclelll MPD detectors was also con~ed. In non-coin~ir1Pnt mode, MPD
devices according to dirre~cllL embo-limPnt.c of the invention had comparable detection 15 effil~iPn~y and about 50 times lower radioactive background as colll~alcd to conventional devices. Also, the conventional systems are calibrated to about +5% wherein our MPD is calibrated to within + 1% .
Table 2 shows some of the most hl~ol~ll p~u~lelcls, namely detection efficiency and background, for a plurality of commercial gamma counters and two MPD detPctors according 20 to the invention based on NaI(Tl) and CaF2 (l~u), respectively. Table 2 also shows that for an MPD device in an AND mode, when coincidence between two photons emitted by an EC
source is used, the background is further ~ pd as cwlll,al~d to an OR (non-coincident) mode, which leads to considerable increased sensi~ivily. However, some reduction of detection efficiency is observed in the coinrid~Pnt mode.

Table 2 Detector Detector EfficiencyBackground for Il2s (cpm) Gamma-counter (1) 70% 200 Gamma-counter (2) 80% 150 Gamma-counter (3) 50% 60 30Gamma-counter (4) 50% 60 CA 0223629~ 1998-04-29 MPDl[NaI(Tl)] (5) 60% (10%) 2 (0.1 cpd) MPD2[CaF2(Eu)](6) 50% (5%) 1.5 (0.3 cph) (1) Rec~m~n 5500 at American Red Cross, Gaithersburg, Md.;
(2) Bec~m~n 5500 Gamma Counter at Lehigh University, Bethelheim, Pa.;
(3) ~J~mm~cope L~B 1272 at Geo~t;towll University, Wasl~in~lon D.C.;
4) ~T~mm~ccope LKB 1292 at MSKCC, New York, NY;
(5) MPD based on two 2" NaI(Tl) scintill~tors;
(6) MPD based on two 2" CaF2(Eu) scintillators.
For the MPD detectors, ~ clroll~lallce is given in both OR and AND COUll~ g modes 10 (values in parenth~ses are the AND mode values).
O~ iOn of MPD systems permitting both OR and AND modes of operation according to the present invention will now be ~ cllcse(l. Sandwich detectors allow a coincident (AND) mode of operation, thus dr~m~tir~lly reducing the background by orders of magnitllde. However, the detection efficiency for this ~q~ ition mode is relatively low (5 to 15% for Il2s sources and 2" to 3" tli~ el detectors in a sandwich geometry) which considerably extends counting times for low-activity sources. However, sandwich detectors according to the invention can also operate in a non-coincident mode, which is referred to as an OR mode. In this OR mode, an event registered in either of the detectors is counted.
Thus, the whole system works as one detector, similar in this respect to a well detector or any 20 non-segm,o.nted detector. In the OR mode, the detection efficiency is higher (typically, 50%
for Il2s sources and 2" ~ m~ter detectors in a sandwich geometry), but the additional benefits of better background rejection through coincidence are lost.
An MPD in accor~allce with the invention operated in the OR mode achieves considerable background reduction through the use of: optimal scintillator thi~lrn~ss; correct 25 inlelpleL~lion of a fraction of two-photon events (50% for a two-detector system) which allows reducing the counting region of interest to the single-photon peak; anticoinri~lenre.; and pulse shape analysis.
In a 2" ~ mloter CaF2(Eu) crystal MPD the background in the OR mode is 1.5 to 2 cpm and the detection efficiency is 50%. During the OR mode ~cqlli.~ition, coincident events 30 are identified and counted, thus avoiding the need for sepal~t~ AND mode counting. Upon CA 0223629~ 1998-04-29 completion of the ~q~ ifion~ the MPD outputs both OR and AND counting data. The OR
data should be used if the activity of the source is above the OR background equivalent activity (about 5 picoCurie, or 2 attomole of Il~s label). These data have a lower st~ti.ctir~l ullcc~ y due to the higher detection efficiency of OR counting. If the OR counts are close to the OR
mode background, the AND mode counting data is used, due to its much lower background.
As a rule of thumb, the OR mode of operation is better for sources larger than 10 dpm, i.e., about S picoCurie. However, in the activity range of 1-20 dpm both OR and AND
counting modes are beneficially used. For this range of sensiLivily, software is used combines the OR and AND data to obtain the best estim~tors of the true count rate. This permits 10 minimi7.ing the artifacts due to st~ti~tic~l unce. ahlly (AND data) and high backgloul~d (OR
data). Below 1 dpm, the AND mode gives a re~on~l-ly better signal to background ratio.
One of the important software functions according to the invention is to ~lu~t:lly estim~te the dead-time and pile-up corrections to the counting rate. The DSO-based acqllicition of pulses involves a relatively large amount of dead time, which has to be 15 co-l-pen~ed for, in particular, for high count rates. In MPD devices according to the present invention, this is done using an additional counter/timer card.
The counter/timer card preferably has at least two pulse counters and a timer. One counter should count all hdldwdie triggers formed by the signal conditioning/triggering card, and the other should count coincident triggers only. The timer should keep track of the 20 exposed time with good precision. In the plefe~led implPmPnt~tion, model PCL-720, distributed by JDR Microdevices, is used. This card has 3 counters, one of which can be converted into a timer by internal wiring. The counters are 16-bit, so they should be read out at least once a second to prevent loss of data. The third counter is wired to count time in intervals of 1/2,500 of a second in this embo~limPnt When counting is initi~t~Pd, the MPD software arms the DSO card for acqlli.cition of the first pulse and simnlt~nPously initiates the counters and timer. Every time the PC
hardware timer hll~llupl comes (e.g., every 55 milli~econds) the values of the counters and the timer are read out and added to the total counts/time elapsed. Upon completion of the counting, the count rate in the applopliate counter (total counts divided by the time read from 30 the timer; the coincident trigger counter is used if the DSO is triggered by coincident triggers, the total counts counter otherwise) is used to adjust the count rates of all events, non-rejected events and events in all regions of interest (ROI's) using the formula:
Cpm; Adj = Cpmj *(CPmTotal Counters/CPmTotal DSO) ~

CA 0223629~ 1998-04-29 WO 97/16746 PCT~US96/16968 where Cpmi Adj iS the count rate (per minute) for the ith ROI, adjusted for dead time losses, Cpmj is the raw DSO cpm for the ith ROI, CpmTO,a, Coun~ers iS the total cpm in the counter, and Cpm Tot.al DSO iS the total count rate before rejection in the DSO.
This adju~tmPnt allows full co~ ensalion for dead time losses, and errec~ively makes the linearity of count to be limited only by the pulse pile-up at high count rates. The self-calibration program permits concordance of the OR and AND counting rate data to, for example, within less than three percent.
Fe.rol,llance charact~ri~tirs of CGX detectors will now be tli~cll~.sed D~LC~lO1according to the invention have radically improved perfo~ e when coll,~dred to 10 conventional devices. The improvements include sel~siLivily, reproduceability, and dynamic range.
Sensitivity: A direct indicatQr of the sel~ilivily (or limit of detection) of a radiation counter is its background equivalent activity (BEA), i.e., the activity of a source which would produce a count rate equal to the background in the detector. This figure of 15 merit accounts for both the background and the detection errlciell.;y of a counter.
The typical BEA for MPD detectors in accordance with the presen invention in an OR
mode (non-coinrj~lPnt detection) is in the range of 3 to 4 decays per minute, which is equivalent to less than 2 picoCurie of Il25. This is based on a detection efficiency (DE) in the OR mode of approximately 50% and a background count rate in the Il2s energy region of 20 interest of 1.5 to 2 cpm. Thus, a 5 picoCurie sample will have a S/B of about 3.
The typical BEA for MPD detectors according to the present invention in an AND
mode (coincidence detection) is in the range of 1 decay per day, which is equivalent to a few femtoCurie. This is based on a DE in the AND mode of about 7% and a background count rate in t_e Il25 region of interest of 1 count per two weeks.
For a small activity source (10 picoCurie), cc,llllll~.cially available detectors are either unable to ~lu~ t~ the sample or provide l~ h~l measurement with S/B close to one.
However, a 10 picoCurie Il25 sample was repeatedly measured with a CGX detector according to the invention, 400 times over a few weeks period. The measured activities are compAtihle with the known half-life of Il2s. These mcas.ll~"llents were obtained using the OR mode of 30 operation, in which coincidence is not employed.
In an enh~nred pclrollllallce mode, which is based on coincidence, and more ~LlhlgellL
pulse shape analysis, the DE is solllcwllaL lower (5-10% vs. 50%), while the background is lower by a few orders of m~gnit~lde. For this el-hAI-red mode, the pelrolllla~lce depends on CA 0223629~ 1998-04-29 W O 97/16746 PCTrUS96/16968 the sample size. For sl~dal.l 12 mm tli~m~oter sample tubes, the DE is 6.5% and the background is 0.25 cph, yielding a BEA of 4 dph. For small s~mrle~ (4 mm in ~ ~r or less), the DE is 5-7% and the bac~l~ulld is 0.5-1 count per week which is equivalent to a BEA of 3 dpd (decays per day). In this enh~nre~l sel~ilivi~y mode it is thus possible to detect Il25 sources with activities of about 10 dpd, i.e., cont~ining less than a thousand Il25 atoms.
Reproducibility: Exceptional reproducibility of measulelllell~s with MPD devicesaccording to the invention has been achieved by selecting electronic e1em~nts with very low lelll~clalulc coeffirient~; partial colllpellsalion of scintillator [CaF2(Eu)] lelllpclalulc response;
rejection of dark pulses due to cosmic rays, and rejection of elecllu...~gnrlir iulelrl~l'cllces.
All these effects are known to be time-dep~n-lent For example, the tellll,elal~
dPpen~lenre of the signal from scintillators leads to day/night effects on the level of 3-5% in conventional colllll~l.;ial gamma detectors. At low count-rates the diurnal and annual modulation of the flux of cosmic rays leads to a few percent effects in well detectors. Finally, sensitivity to elecll~ agnptir illl~lr~lcllce leads, at activities below 10 cpm, to noticeable day/night changes. The rejection/co--.l~en~-lion of these effects in the MPD considerably improved measulclllent reproducibility and removed all diurnal effects.
The mea~ulclllclll reproducibility of MPD devices according to the invention is del~. -.-;.-~d primarily by counting st~ti~tirS~ clock accuracy, and the reproducibility of placing the sample within the instrument. MPD devices according to the invention are reasonably stable over the long term. No variations in background have been observed, and the detection efficiency is stable. To test the measurement stability of an MPD device over a few weeks, the same sample (50 nanoCurie) was counted about 1,200 times, removing and replacing it in the holder before each measurement. For each measurement the sample was counted until 10,000 counts were ~rcllmnl~t-qd (equivalent to a st~ti~tir~l uncertainty of +1%).
Another advantage of the MPD according to the present invention is the possibility of reliable calibration. Improved calibration is possible due to use of the "sandwich" geometry which ~imini~hr~ absorption and sample pl~remrnt artifacts, and the use of Il25, pe- .-~ g use of the Eldridge calibration procedure. For example, the same MPD device was calibrated 25 times with a 50 nanoCurie source. The average DE was delellllined to be 49.1% with a standard deviation of + 1. 7 % . In comparison, the typical commercially available detectors are nominally calibrated to within _5% but in reality for sources with activity below 0.1 nanoCurie, the calibration uncertainty is closer to _10%.

wo 97/16746 PcTruss6/l6968 D~namic range: Linearity over many orders of m~gnih~rle is a desirable feature for all analytical and biom~ ~liral applications. Often, the levels of primary biocompound should be compared with the level of metabolites, which can be many orders of m~niblde lower. The majority of ~;ullcnLly used conventional illsllull~ s have very limited dynamic 5 range.
Photographic emulsions permit y~ i\re measurements only over 1.5 logs range.
Typically, the response of the detectors is limited both at low levels and at high counting rates.
Tnus, instead of the desired linear l~ ,onse, the characteristic detector's respollse is an S-shaped curve. At low count rates, well detectors are seriously limited by the intrinsic 10 background. Similarly, optical detectors, e.g., color ~e.,~loll~eters, are severely limited by photonic background. At high activity levels, pile-up or optical il~t~lÇ~,r~llce distorts the lhlealily of conventional detectors. Actually, gamma ~leleclo,., are close to ideal for high count rates, up to about 1,000,000 cps. Thus, typical well counters show a linear response over about 4 logs dynamic range, from about 100 cps to 1,000,000 cps.
The background rejection teçhni-lues used in MPD devices acco~hlg to the p~sse invention permit reliable qll~ntit~tion from 1 cpm to 1,000,000. cpm, i. e., over 6 logs dynamic range in the OR mode. When using the e~,h~n~ed operation AND mode, for which thebackground is 1 cpd, the MPD detectors according to the present invention are linear over 9 logs dynamic range. At higher count rates (above 500,000 dpm), saturation may be caused 20 by pulse pile-up in the scintillator. Although the response is no longer linear in this range, the dead time of the MPD is non-exten~l~ble, so that counting can be ~rolllled and the results corrected for pile-up. At low count rates, the linearity of response is limited by the background. The results of measule~ n~ using MPD are shown in Figure 4. Dilutions of Il25-labeled reagents show perfect linearity of measured activity over 5 orders of m~gnitllfle 25 down to the 0.1 zeptomole/sample level.
The linearity of response of the MPD according to the present invention was compared with comrnercially available colorimetric detectors and the advantage of the MPD is evident.
The dynamic range of the MPD detector is considerably better than for color ~e~ ,llleters.
To perforrn this study, streptavidin-HRP was io-lin~ted and the same sample was measured 30 using MPD and a commercial color spectrometer used for ELISA tests. The results are plcsenl~d in Figure 5. The sensitivity of the MPD is at least two orders of m~gnitllde better than that of the color spectrometer, and its dynarnic range is about five orders of m~gnitllrle CA 0223629~ 1998-04-29 W O97/16746 PCT~US96/16968 better. In the range where the color spectrometers work reliably, identical results were obtained using the MPD according to the present invellloll and the color spectrometers.
Use of CaF2(Eu) scintillators is now ~li.cc~ ed. NaI(Tl) scintillator crystals seem a natural choice for MPD instruments. In coinri~lPnt mode, background rejection is inversely 5 plupolliorJal to the square of the energy resolution, which for NaI(Tl) is about 50% better than for other scintillators. Fullll~llllol~, among scintillators with reasonable energy resolution (NaI(Tl), CsI(Tl) and CaF2(Eu)), sodium iodide scintillators are fastest. According to ~ d~'d practice, background rejection is proportional to the square of the timing resolution. F.~tim~t~s suggest that the background in an MPD based on NaI(Tl) should be about four times lower 10 than when using other scintill~tors, and such an MPD system for very small, say < 10 microliter sources, achieved a background of about 0.5 count per week (0.5 cpw). However, disadvantages of NaI(Tl) based devices include:
* NaI(Tl) is mech~nir~lly fragile, e.g., it often cracks when submitted to telllpel~lule gradients and/or during transportation;
15 * NaI(Tl) must be h~rm~tir~lly sealed, and the thin Al or Be foil covering the front surface of the scintillator is easy to rip off when samples are placed in its vicinity, e.g., when opelalillg spatially resolving MPD devices;
* When larger samples are used, the backgr~ul~d in MPD ~y5l~;llls based on NaI(Tl) significantly deteriorates due to X-ray cross-talk b~,~weell the crystals.
Replacing NaI(Tl) with other scintillators, inrl~rlin~ CaF2 (Eu), leads to a small increase of background. However, by opli,.,i~i"g read-out electronics and data proces.cing software, for small samples of Il25 or I'23, CaF2(Eu) based MPD systems reach almost the same background as NaI(Tl) based systems. For large samples, the bac~ruulld in CaF2(Eu) based MPD systems is about a factor of ten better.
CaF2(Eu) has the above mentioned surprisingly low background for Il25 detection for several coulll~li.l~uitive reasons. First, for MPD systems according to the invention, the main source of background in the AND mode is the detection of a soft X-ray emitted and absorbed in one crystal coincident with some source of energy detect~d in the second crystal. When NaI(Tl) is used, any absorbtion of an external photon with E > 35 keV in the crystal leads to - 30 remission of 26 keV or 32 keV photons from rearrangement of atomic shells. Thus, characteristic iodine X-rays are emitted when the NaI(Tl) crystal is used. These cannot be - distinguished from the 25 and 31 keV Te X-rays emitted by either I'23 or Il25 t~ ghter nucleii.
Actually, for CsI(Tl) this effect is even bigger because the characteristic X-rays of both Cs and CA 0223629~ l998-04-29 W O97/16746 PCTrUS96/16968 I are virtually the same as X-rays emitted by radioiodine. Fo. ~unalcly, CaF2(Eu) on the other hand includes only low atomic number elements. Thus, its characteristic X-rays have less than 15 keV energy and can be dirrc.cn~;~t~cl from radioiodine X-rays. Thus, preferably NaI(TI) or CsI(Eu) should be used for EC radioisotopes with atomic number of either less than 40 or larger than 70. For EC isotopes with atomic number between 40 and 70, CaF2 (Eu) is a plcr~ cd scintill~tor.
A second advantage of CaF2(Eu) is even more subtle and coùlll~lil.luilivc. CaF2(Eu) is a very slow scintillator with characteristic light decay of about 5 mic.~,seconds, i.e., about a factor of 25 slower than NaI(TI). Conventional detectors used the fastest possible ~l~tectors, 10 with backgrounds of about 1 cps. When background is pushed to a few cpm, the do~ illg sources of background are due to cosmic rays and dark current pulses from the PMT's.
Spurious signals due to energy deposited by cosmic rays in the crystal itself can be partially accounted for by analyzing the amount of energy deposited. Particularly for low energy X-rays, e.g., for Il25, this method permits rejection of over 95% of cosmic rays crossin,e 15 sr,intill~tors. This includes efficient rejection of secondary particles in the cosmic ray showers.

However, in the case of an MPD system, the crystals themselves are very thin and the surface of the PMT cathodes is about fifty times larger than the surface of the crystals. A
high energy cosmic ray striking one of the PMT anodes gives rise to an avalanche of electrons which are subseqllently amplified. Such pulses lead to an al,pa-enl energy deposition much 20 lower than the energy of the cosmic ray, i.e., there is a considerable overlap bclweel1 the energy ~l,ecl ul-- of cosmic ray in~ r,ed dark current pulses in the PMT's and the energy from radioiodine. In detectors using scintillator-PMT combinations this source of background accounts for a few counts per minute in each PMT, or a few counts per hour in the coincidence mode. However, the energy deposited in the scintill~tor typically leads to pulses 25 longer than the cosmic inr1uce~1 pulses in the PMT, which have a chal~ le. ;~l ir time constant of about 0.2 nsec.
In practice, the rejection capability is limited by several detector char~cteri~tirs. These include the characteristic ~c~ollse time of the scintillators, the parameters of the shaping amplifier, and the performance of the on-line pulse shape analysis system. For example, in 30 MPD systems there is a trade off bclween the need for low cost and the pclrollallce of the - on-line digital storage oscilloscope (DSO) used in pulse shape analyses. Only about half of the pulses in~luced in PMT's by cosmic rays can be rejected versus pulses inriuce(l in NaI(Tl).

CA 0223629~ 1998-04-29 WO97/16746 PCT~US96/16968 The dirr~ ce in pulse rise times btLweell cosmic ray inrluced PMT pulses and those created in CaF2(Eu) is large, i.e., 0.1 nsec and a few microseconds respectively. In a CaF2(Eu) based MPD system, over 95% of cosmic ray in(1llre-l PMT artifacts are rejected on-line.
Changes were obeserved in NaI(Tl) crystal ylop~llies occurred over a 2 year period in about 40 NaI(Tl) crystals. Some (about 10%) crack from thermal stresses while others (about 15%) turn yellow due to their hygroscopic prui)elly. In contrast, no ~ignifir~nt variability in crystal properties was observed for twenty CaF2(Eu) crystals over a one year period.
In ~ a~ CaF2 (Eu) scintill~tQrs in sandwich geometry permit achiev~lllellL of 10 excellent background rejection, especially when large rli~m.-ter samples of radioiodine are measured. The excellent m~ch~nir~l pl~tlLies of CaF2(Eu) provide another advantage over NaI(TI).
Selection of scintillators and their ~lim~n~ions will now be described. Conventional scintill~tQrs are ul~Limi ed to m~ximi7e the detection efficiency. Thus, the typical thirknPs~
15 select~d is about twice the stopping power at the energy of interest. Also for typical sample sizes, say 0.5 inch in tli~mPter, the ~ m~ter of the selected crystal is 3 or even 4 inches.
However, background is roughly proportional to crystal volume in a complicated nonlinear function of scintill~tor dimensions. For example, for 3" crystals conventional designs couple them to 3" PMT's. However, 3" inch PMT's are considerably more radioactive than 2"
20 PMT' s. This radioactivity originates from naturally occurring isotopes in the PMT glass. The larger surface area and greater thir~nrss of the glass walls in larger PMT's thus leads to significantly higher radioactive background. Thus, against the conventional as~wllyLions, to .e the performance of low background MPD devices, 2" di~mrter detectors and subst~nti~lly thinner crystals are preferred in the present invention.
MPD detectors according to the invention have the advantage that they can be self-calibrated for I'2s using two photon coincident detection. However, to accomplish this, the pair of detectors should be well m~tt~h~d, i.e., should have similar proyelLies. To facilitate the selection of well m~tch~d pairs, all crystals within a given MPD system are preferably cut from the same large di~m~ter crystal. The crystals are mounted in low radioactive background 30 copper tubes. The X-ray window/reflector is made of MgO and 50 micron thick Al film.
Optionally, thin teflon film is used. The optical window is made of at least 2 mm thick - quartz.

CA 0223629~ 1998-04-29 Optimization of diameter and thickness: For single-sample MPD devices, accordingto the invention the o~lilllulll scintillator size was found to be 2" rii~mPter. For this size the S/B is highest for the standard sandwich geometry, i.e., when a sample is placed belwee detectors spaced about 1/2" apart. However, the o~ lli~lion curves as a function of crystal 5 size are rather flat. For example, (S/B)[3"] ~ 1.2. When op~ .i..g MPD detectors for Il25, the oplilllulll thirlfn~ss of the scintillator is 1.5 mm for CaF2(Eu); a change of crystal thirl~n.osc from 1.0 to 3.0 mm changes the (S/B) by only about 50%.
Selection criteria: Preferably, each crystal is optically inspected; crystals which are cracked or not p~lr~;lly L~ nl are rejected. The crystals are then mounted on select~cl 10 low radioactive bac~roulld PMT's and are submitted to a series of acceptance tests. The first test checks for the energy resolution and the detection efficiency of the whole crystal.
Unfullul~ly, PMT's are far from being unirollll, and the performance of dirr~l~,.ll zones of the PMT should be measured. To do this, the surface of the scintillator/PMT assembly is chi~lded by rings of lead of increasing diameter with a ra~iioactive source placed in the center.
15 The first lead mask has a 0.5 cm hole. The energy resolution in the center of the scintillator/PMT is then ch~c~d Two other lead masks are also used to check the energy resolution, a one inch and a two inch lead ring. Only CaF2(Eu) crystals with dE/E(FWHM) 21% for Il25 are accepted. Furthermore their radioactive background must preferably be ~ 0.05 dpm. Typically two to three crystals from each batch of ten are rejected and returned 20 to the producer.
Selection of low bac~luulld PMT's and optically ll~l~s~ ll material between the PMT and scintillator crystal is now described. MPD devices according to the invention require considerable care in the selection of PMT's. Custom made quartz PMT's are ~ler~lled if the additional cost is tolerable. Off-the-shelf integral scintillator/PMT assemblies 25 and commercially available PMT bases may be used instead, with a~ropliate modifications in accordance with the invention. A large part of the r~lioaçtive background is due to the use of st~inles.c steel in these integral assemblies, and commercially available PMT bases are quite radioactive. PMT bases according to the invention have a composite shield between the base and the PMT to tlimini.ch the radioactive background. Also, the PMT's are optically coupled 30 to the scintillators via an appropliate low radioactive background optical coupler (quartz).
To ~liminich radioactive background in MPD systems, PMT's are selected which areoptimal for both 2" and 3" scintillation crystals. The following PMT pal~ll~lel~ are of importance: radioactive background, detection efficiency, energy resolution, homogeneity over CA 0223629~ 1998-04-29 WO 97/16746 PCTAUS96tl6968 the photocathode surface, dark current, long term stability, and depen~enre of signal on clll~lallllc and m~nPtir field.
Testing procedure and results: C~n~ tP photomultipliers from five dirrclcn m~mlf~rtllrers were tested, with the best results obtained for Electron Tubes, Inc. (ETI) and 5 H~m~m~t.~u PMT's. About 40 PMT's from these two ~ -r~cl~ were tested for theirelecLrol~ic prollcllies and radioactive co--l;..--in~lion. The H~ PMT's provide marginally better electronic pc,ro"lia,~ce. They feature both an excellent detection efficiency and good energy resolution. Also, their dark current is somewhat lower. However, for both 2" and 3" tubes, the best individual tubes were from ETI. These tubes have a somewhat larger distribution of elccLlollic pelÇo""a~ce within the batch. COllC~ g the radioactive background, the ETI tubes are preferred; measured bac~,oul~ds were a factor of ten lower than for H~m~m~t~u tubes, for ex~ll~lc.
The PMT's are subjected to a series of tests to deLe.,llille whether they meet the selection criteria. The first test checks for energy resolution and detection efficiency on an open faced crystal. Next, the energy resolution for the different parts of the PMT is c~ PC~P-d.
This is done by using three dirr,lc,lt lead apelLu~cs, in which a radioactive sample is placed.
The first mask has a 0.5 cm hole and the second mask is a one inch outer lead ring.
Electronic palalllclcl~ are strongly dependent on the energy of the isotope used. For example, there is no correlation found between the energy resolution at about 30 keV (Il2s source) and at about 88 keV (Ga67 source).
The relative radioactive bacl~,vu"d in the PMT's is measured by placing a PMT next to a 2" diameter, 2 mm thick CaF2(Eu) detector coupled to a selected low-background 2"
PMT. The whole test system is well shielded with lead, tin and copper. The background in the CaF2(Eu) detector is measured in the energy range of 20 to 40 keV (the energy region of interest for I'25) and cG",~ared with the background without the studied PMT. Pulse shape analysis is used to discrimin~te between events and electronic artifacts. Dining testing the background in the CaF2(Eu) detector in the absence of the test PMT was found to be 0.7 cpm.
ETI PMT's normally increased the count rate only in.~ignifir~ntly, while the H~
PMT's generally created a large additional backgl-ulld.
The radioactive background found in the H~m~m~t~11 PMT's is ~ullllisingly large; the average background is 3.5 cpm but values as high as 4.2 cpm were observed. Thus, selected - 2" ETI Model 9266KB PMT's coupled to a NaI(Tl) or CaF (Eu) scintillator are plcfcll~,d.
The average dE/E(FWHM) = 17.4% and the ",ini~lm~ and m~xim-lm energy resolution is CA 0223629~ 1998-04-29 W O 97t16746 PCTAJS96/16968 16.2 % and 20.7 %, respectively. The average detection efficiency is 37.4 % with ~--;.-;..-~.-and m~ximnm values of 31.1 % and 40.3 %, respectively. The radioactive background due to the EMI PMT's is rather low; the average background is 0.2 cpm with ...;n;...~... and m~ximllm values of 0.1 cpm and 0.55 cpm, ~c~ecli~ely. About 20 % of PMT's were 5 observed to have backgrounds higher than 0.3 cpm. After these were rejected, the average background is 0.13 cpm with minimllm and m~ximllm values of 0.11 cpm and 0.28 cpm, ~speclively. Another plcrellcd model is ETI model R-2486 PMT.
Pacl~gin~: Conventional PMT's are typically p~c~ged in prefabricated ~ll.. ;.. ~
or st~inles~ steel tubings. These commercially mounted PMT's have increased radioactive 10 background and so a modified mounting is ~lcfellcd. PMT's shall be rh~r~Pcl for cracks or flaws. The PMT is washed with isopropyl alcohol to remove any particles or co~ -in~ c.
Next, the glass surface with the exception of the window area is covered with four layers of black electrical tape. After this is complete, one spiral of black electrir~l tape covers all the bands except for the first (next to the window). Subsequently, two layers of copper foil tape 15 with conductive adhesive cover the whole PMT, inr.lu~lin~ the plastic base. Another spiral layer of black electrical tape covers the copper. Next, a single layer of copper foil tape followed by a single spiral layer of black tape completes the Wl~illg process.
The crystal is coupled to the PMT with silicon optical grease. Care is taken not to introduce air bubbles between the crystal and PMT. Next, white Teflon tape is wrapped 20 around the PMT where the crystal and the PMT meet. The crystal is held in place with four to eight strips of very thin adhesive ~ll....i...,... foil. A layer of adhesive copper tape is placed around the crystal and PMT to secure the PMT and crystal together, while providing protection against extern~l X-rays. The three inch PMT's have adhesive lead tape secllring the two together. Finally, to ensure against visible photon leaks, a spiral layer of black 25 electrical tape covers the copper.
For low-background counting the scintillators should be shielded from radioactivity in the photomultipliers to which they are coupled. It is impossible to shield the detectors from high-energy gamma rays from the PMT's without degrading the optical properties of the scintillator/PMT system. It is possible, however, to shield the scintillators from beta particles 30 and low energy photons by using a Lldl~dlenl window such as quartz between the PMT and the scintillator. This window 24 is shown in the exemplary embodiment of Figure 1, with - optical grease layer 23. Quartz was select~d both for its excellent optical plol-cl~ies and its high purity. Quartz m~tch~s the optical density of CaF2(Eu) very well and is acceptable for CA 0223629~ l998-04-29 NaI(Tl). 5 mm thick quartz windows are plc~ d. There is no observable radioactive co~ ion in quartz.
Improved ~hiPl(ling of the scintillators from gamma and X-rays can be achieved using materials with higher ~Lo~ lg power than quartz. High-purity GeO2 and g~....~.i...ll-based 5 glasses are plefel~d for this purpose, for they have very low intrinsic radioactive background.
Their higher atomic number and density is an advantage over quartz windows. Such windows with a few mill ;l l~ h;rL I IP~ efficiently stop low energy photons, as well as beta particles, without degrading the optical qll~litiPs of the scintillator/PMT systems. The optical plop~,llies of gelica and ge---~ glass match NaI(Tl) better than does quartz.
Another alternative is to use high density glasses based on lead, particularly high density ~la~ lL crystals such as PbF2 and bisllluLh g~ lAllile (BGO). In the case of BGO, undoped crystals should be used so as not to gel~la~ artifacts due to scintillAtit)n within the BGO. The optical density of these materials is higher than for CaF2(Eu) or NaI(Tl). Thus, it is ~lefell~d to use a thin layer of special optical greases, e.g., powdered PbF2 in silicon 15 grease, to match the optical pro~ellies of the scintillator and window, and the window and PMT. Optionally, a window col~i~L,llg of a triple sa~lwich consisting of gelica/high density window/gelica can be used. The thir~nP~s of the gelica may be much smaller than the high density optical window.
Shield/Sepal~l~Jl. FxtPrnAI shielding of an MPD according to the invention is required 20 to stop ambient radiation, as shown in the exemplary embodiment of Figure 1 as shield 55.
A 2" thick Pb shield is ~ Pqll~AtP for MPD detectors built around 1-1.5 mm thick scintill~tnr crystals. Increasing the shield thir~nPc.c further does not signifir~ntly reduce the background on the Earth's surface, most probably because the rem~ining background is predo~ ly due to cosmic rays. However, if the shielding is made of lead only, secondary lead X-rays are 25 present in the background spectra due to excitations in the shield itself by radioactive particles.
Composite shielding is thus plefe"~d, with a 1-5 mm thick layer of tin inside the Pb shield to absorb the lead X-rays and a 1-5 mm thick copper layer inside the tin layer to absorb the tin X-rays. The X-rays of the copper itself are of a sufficiently low energy (8 to 9 keV) to be outside the Il2s region of interest. Both NaI(Tl) and CaF2(Eu) scintillators have sufficient 30 energy resolution to reject these pulses with better than 90% probability. Commercially available Cu foils are sufficiently pure and do not introduce additional radioactive background.
- The external shield should enclose the detector assemblies, including the PMT's and bases, on all sides.

CA 0223629~ 1998-04-29 WO 97/16746 PCTrUS96/16968 Sample holder/crosstalk eli,-,i,l~lol~ are also made of copper, lead, or composite copper/lead plates 1-5 mm thick. This thir~nPss is sufficient to stop iodine X-rays.
The materials for the shield should be tested for the absence of r~ o~rlivily~ by mP~lring the background in a CaF2(Eu) detector selected for low radioactive background and 5 placed in a selected, low background shield. A test p~lro~ ed for 2 hours yields a ~L~t~ r~l ullc~ lly for each point of about + 10%. Each shield which has a r~lioactive background larger than 0.9 cpm may be remPa~l~red for 4 hours. If the new value is still above 0.9 cpm the shield is rejected. Typically, 20% of shields have to be rejected.
Integral PMT base assembly for high stability MPD devices is now described.
10 Accurate counting of low activity sources requires a high stability detector system. The observed drifts are predominantly due to chdnges in ambient l~ lulc, which cause drifts in the yield of the scintillators as well as the gains of the PMT's and the electronic readout.
Improved electronics and high voltage power supply are p~relled for stabilizing the PMT's.
The high voltage power supplies (HVPS) for the PMT's should be stabilized by 15 introducing a high gain negative feedb~cl~, using as a ref.,lcl~e chips (e.g., Max 580) with low telll~)clalule coefficient (1-2 ppm/degree Celsius), and using 1% metal film resistors with a low l~l,lpclalu,e coefficient. The voltage divider for the PMT is based on the same type of l~Si~tOl~. The t~",~e,alulc depen~lPIlr-e of the preamplifiers is Illil~illli7Pd by using highly stable op-amps and distributing the gain so that each amplifier cascade has a gain lower than 20 5.
The problem of long-term and temperature-in-i--ced drifts in the PMT's and scintillators can be compçl-.c~Pd for by adjusting the gains of the amplifier, so that the acquired spectra are ~ rh~.~ged. Electronics permits digitally controlling these gains. When coupled with telllpclalule readout devices placed in the detector system this allows continuously adjusting 25 the gains to compensate for tcllll)cldLul~ in~nr~ed drifts. The tcl~,~elalule-to-gain calibration for each detector can be efficiently obtained by a series of measu,elllcll~ in non-coincident (OR) mode and used for both OR and AND mode acquisition. The entire calibration procedure can be automated and p.,lrol,lled periodically to colllpensàl~ for long-term drifts.
PMT Bases: Commercially available HVPS and PMT bases are not ~iP,qll~te for low 30 background detectors for biomP-~lic~l applications due to depenrl~pnre of HVPS and voltage divider (VD), perfol " .~ . ,re/risk of high voltage cable, radioactive background of elements and ele~;Llo",~gnPtir pick-up noise b~lweell the PMT and preamplifier. Thus, an integral PMT
base is ~lcr~ d, cont~ining a HVPS/VD/~ lp/shaping amplifier assembly.

CA 0223629~ 1998-04-29 wo 97/16746 PCT/Us96/16968 Convention~l PMT bases use much less integrated designs. Cl~ c~lly, only a voltage divider is placed in the PMT base. T_is leads to considerable electronic pick-up noise that is undesirable in the MPD. Several c-)-,-.n~r~;ial devices feature PMT bases with integral voltage divider and preamplifier. For m~xim~l integration of t_e PMT base, it is l~refe~ d to include 5 the HVPS, voltage divider and preamplifier because the use of high voltage cables leads to problems with ground loops and eleclro--.~gn~tir pick-up and may pose the risk of elecLlv~;ulion, especially with portable devices when used in high hllmitlity envho~ ellLs.
The r~1ioar-tive bac~lvulld of PMT bases was measured by placing the PMT next toa 2" ~ mPter~ 2 mm thick CaF2(Eu) detector coupled to a se!ecte~l low-background 2" PMT.
10 The whole system was well ~hi~l~ed with lead, tin and copper. Then t_e background in the CaF2(Eu) detector was measured in the energy range of 20 to 40 keV (the region of interest for I'25), and colll~ed with the background without the studied PMT base. Pulse-shape analysis was pclrolllled to discl U~il~t~ b~Lw~ell events and elecLrollic artifacts. The bacl~l~ulld due to PMT bases was measured with bases 2 mm and 5 cm from the surface of 15 the CaF2(Eu) crystal. The first positioning permits measurement of the bac~rvulld while the ,lea~ul~lllent at the second position in-lirat~s how much this infl~enres the MPD perfonn~nre (length of used PMT's is about 5 cm). The background in the CaF2(Eu) detector in the absence of test PMT bases was found to be 0.7 cpm.
When placed 5 cm from the scintillator, the background due to the PMT base assembly 20 does not inflll~nre overall background of the MPD. However, it is sul~lisingly large when placed close to the scintillator. Thus, several PMT bases were ~ s~mhled and theradioactive background of individual components, was tested, namely:
* plastic HV connector;
* divider chain resistors and capacitors;
25 * Pb/Sn/Cu composite shield;
* ~ .";".,.., holder and h~ in~ of PMT base;
* HVPS module and preamplifier module.
Sul~lisingly, it was found that the largest radioactive background is from the plastic high voltage connector, mostly due to beta particles. This bac~gloulld can be very efficiently 30 ~ttPml~t~.~ by a t_in plastic guard ring. After this modification, the PMT base radioactive backg,v~ d accounts for less than 10% of radioactive background of PMT's.
- Qll~ntit~tive measu~ ent of low activity sources, say ~ 0.1 picoCurie, may require measurement times of up to several hours. Any slow drift of electronics can therefore CA 0223629s 1998-04-29 W O 97tl6746 PCTAJS96/16968 inflllrnre these long term (1 hour) mea~ulcllRllL~. The main source of this drift is the PMT's.
The strong depçn~1~nre of the amplitude gain on the high voltage from the HVPS leads to lulc-dependent signal drifts.
High voltage power supply: The voltage divider supplies the negative voltage from the 5 HVPS to the PMT dynods (pins 1-11) and to the cathode C via an RC circuit. This is shown in Figures 3A, 3B, and 3C in PMT base assembly 30. The voltage divider resistorscsclllcd as R7 and R8 have a low Iclllpc,~lulc coefficient (~ 100 ppm/~C) and thus stabilize the PMT output signal. Negative HVPS 50 preferably includes a regulated DC to DC
CO~ Ol 64 from M~t~lls~ Co., voltage regulator 66 and a fee~lbacl~ circuit with precision reference 68 (IC1~?e1~IU1C coefficient about 3 ppm/~C) and conll,aldlol amplifier 70. F.l~m.ontc with low lem~cl~lulc coefficient are plcr.,llcd because of the importance of ~limini~h~-l telll~ lulc ~ep~n~l~nre of HVPS output.
Ten-fold better performance can be achieved than when using commercially available NIM modules such as are available from Ortec, Canberra or Tennelec. This ~l;",i"i~l~r(l the 15 diurnal variability of the MPD con~ rrably.
A pler."l.,d HVPS has the following chal~clclislics:
~ Input voltage: -15 to -18 VDC
~ Input current: _ 120 mA
~ Output voltage range: 300-1,100 V [DC]
~ Noise: _ 50 mV peak-to-peak ~ Tclllpe,~lulc stability: ~ 20 mV/~C at nominal output voltage (1,000.
V) Another important element of the PMT base is optimized preamplifier/shaper 52 shown in Figures 3 and 3C. A triangular pulse shape is optimal when using pulse shape analysis to 25 reject background, e.g., due to dark current pulses in PMT's. In conll~-sl, the majority of commercial devices use ~ls~i~n pulse shaping, which oplillli~cs the energy resolution for low energy X-rays and is prcrcllcd when pulse height analysis is pclrolllled~ e.g., using mllltirll~nnrl analyzers. The amplifier/shaper preferably includes a pre-amplifier 52 based on an EL 2030 low-noise current feeclb~ amplifier and an output amplifier 54 based on a model 30 OP64 amplifier with high gain and dynamic range. Output amplifier 54 does not need correction and tuning during pclrollllallce. The pulse dirre.clllialion chain Cl, Rl and integration chain C2, R2, as shown in Figure 3, define the parameters of the output pulse:
~ Rise time tr~ 400 ns [800 ns]

CA 0223629~ 1998-04-29 ~ Pulse width at 0.1 FWHM t = 3 ms [4 ms].
The first numbers correspond to the use of NaI(Tl) scintillators while the numbers in parentheses correspond to CaF2(Eu) scintillators. The resistor chain R4, R5 defines amplifier 54 gain. The gain was chosen in the interval 20-50 and is adjusted by potentiolll.,t~ . This - 5 value depends on the plop~llies of the given PMT and scintillator crystal and provides an output amplitude of about 2 V for 30 keV photons.
The reslllting PMT base/operational amplifier assembly according to the invention has the following properties:
~ Noise C 50 mV peak-to-peak ~ Gain 20-50 ~ Dynamic range 50,000 ~ Offset <2 mV
~ Input signal tr=200 ns, t=100 ms, [negative polarity pulses]
~ Output signal tr=400 ns, t= 3 ms, [positive polarity pulse]
~ Max. output signal 10 V
~ Nonl;.-~. ;ly 5 2% over the full dynamic range ~ Power source DC current <20 mA, Voltage from +12 V DC up to ~18VDC
~ PMT base size 2.2" ~ ."Ptel, 3.75" length ~ Weight 1 lb (this includes 0.5 lb of internal Pb/Cu shield) These electronics have high temperature stability and do not require adjlls~m~nt during operation. The device is also easy to produce and tune. When compared with NIM modules, they are 10 times more stable and considerably cheaper to produce.
Noise and temperature stability: The high st;llsiliviLy and low bac~loulld laluirelllents of the MPD drive the specifications for the elecllol~ics. Conditions which cause a decrease of spectral sensitivity and S/B ratio can be separated into short-term noise (electronic noise, dark pulses) and long-term drift of electronic pa,~ll-,t~l~ (gain, high voltage, ~ c;l~ule drift).
The energy resolution dE/E(FWHM) at 30keV for plefel~ed ~le~ectors according to the invention is about 16% for NaI(Tl) and about 35% for CaF2(Eu). If electronics are to contribute less than 10% degradation to the energy resolution, this leads to a noise to signal - ratio (N/S) of ~ 0.016. The amplifier, high voltage power supply (HVPS) andelectrom~gn~tic shielding allow obtaining N/S C 0.005. The majority of electronic noise CA 0223629~ 1998-04-29 wo 97/16746 PcT/uss6/l6s68 sources are filtered by the electronics, while the rest of the noise is rejected by on-line sorl~ale shape analysis.
When using colll,llclcially available HVPS, the te~ )claLule depen-len~e leads to easily measurable drifts of pulse amplitude for I'2s. With prcr~ d HVPS according to the invention, 5 the telllpeldLulc depenf~Pn~e of the gain upon ambient temperature is below measurable accuracy. To decrease the inflll~nre of HVPS Lelll~claLulc variation, the voltage divider for the PMT's can be implçm~nt~od using metalloceramic 1% resistors with a low telll~)eldLulc coefficient. FulLll~,llllore, current feedbac~ and the use of a super stable source of baseline voltage allow for a small size HVPS (see Figure 3) with the following parameters:
~ DC voltage input: -12 to -18 V output: -300 to -1,100 V
~ DC current input: 5 120 mA output: < 1 mA
~ Output noise 5 100 mV
~ Pulse noise 5 150 mV peak-to-peak with 100 kHz frequency For example, the pulse noise is a factor of 2-3 better than for commercially available HVPS.
The telll~lalule depen~l~n~-e of HVPS output voltage bcl~cn 25~C and 70~C was compared for commercial HVPS and the HVPS according to the present invention. The baseline/noise shows a better than 1% stability of the baseline during a 24 hour period (measured in winter with heating switched off, i.e., leading to a day/night variation of about 15~C). The pclro~ e of the negative HVPS according to the invention was compared to 20 HVPS PS1800 series (Electron Tubes, Inc.). Both HVPS's were switched on at the same initial tcllll claLule (25~C) and then were heated simlllt~n~ously up to 60~C. The commercial HVPS showed a few percent drift of voltage, but the output of the HVPS according to the invention demonstrated no ~isc~rn~hle variation due to heating.
Single Sample MPD: A plcfellcd implelllcllL~Lion is now described. Existing gamma 25 counter designs typically optilni;~e (letection efficiency. According to the invention, the goal is to ,..i.~ilni,.o background while keeping detection efficiency reasonably high. To reach this goal it is n~cess~ry to elimin~te sources of radioactive background, employ optimal counter geometry (including of shields and sc~Lor), carefully select components, use stable electronics, and use on-line sorLwale for background signal rejection.

~8 CA 0223629~ 1998-04-29 The MPD ll,e,.,role includes the following sub-~y~lellls:
~ photon detectors, e.g., scintillators with PMT read-out;
~ a Se~dldlor/Shield subsy~
~ read-out elecL~ol~ics; and 5 ~ a data ~cq~ ition/analysis subsy~lelll.
CaF2(Eu) scintillators are ple~ ,d for their good stopping power and reasonable energy resolution as previously described. Such systems achieve exceptional background rejection resllhing in an improved cdyacily to qll~ntit~t~ minute traces of target isotope. The device can be operated in two modes as previously described. In the OR mode we achieved 10 high DE (> 50% for I'2s) is achieved with a radioactive backgl~,u"d of about 1 count per minute (1 cpm). In the coinrident (AND) mode, the DE for I'25 is below 20%, but achieves an ultra-low radioactive background of 1 count per day (1 cpd). These results were achieved at the earth's surface with a relatively small shield (about 20 kg of lead). The system is ~le~ign-od around low cost commercially obtainable co,l,yolle,ll~ and well established 15 téchnologies.
A plefell~d imp]em~nt~tion of the MPD detector oylillli;~ed for detection of I'25 is now described. A block diagram of the MPD detector is shown in Figure 1 and an elevation is shown in Figure 2, including "sandwich" geometry, multiphoton coincidence and pulse shape analysis subsy~.lellls. Detector 20 inrllldes two modules separated by low radioactive 20 background SêydldlOl 32 collsislillg of heavy metals, typically a sandwich con.~icting of a few milli",rlels of Cu/Sn/Pb. Inside SèydldlOl 32, a hole 34 is disposed into which sample 36 is to be placed.
The assembly of the two detector modules 20 is enclosed in a heavy metal composite passive shield 55, built of radioactively pure materials, typically lead, tin and copper as 25 previously described. The highest atomic number material, e.g., lead, is most external and at least 1 inch thick. It is followed by a lower interm~ ry atomic number material, e. g., tin, with a few millim~t~rs, e.g., 1-2 mm, thirkn~ss. Finally, a few mm (typically 2 mrn) layer of very pure copper is used.
Each of the detector modules includes the following elements:
30 * inorganic scintillator crystal 22, with thirkn~s.s optimized for a given emitter;
* high purity optical window 24, placed beLween scintillator and PMT;
- * select~d, low radioactive background PMT 26;
* graded passive shield 46 beLwèen the PMT 26 and PMT base assembly 30;

CA 0223629~ 1998-04-29 WO 97/16746 PCT~US96/16968 * PMT base assembly 30 with of high voltage power supply 50, voltage divider 51 and preamplifier 52 (shown in Figure 3).
CaF2(Eu) crystals less than 3 mm thick are plercllcd as scintillators. Optical window 24 is at least 4 mm thick ultrapure quartz, optically polished and provided with optical coupler 5 23, a low radioactive background silicon grease.
Selected PMT's 26 are made of glass with low co~ ion by K40, preferably 2"
PMT's preferably selected to present less than 0.1 cps background. It is preferable to decouple the PMT base 30 from the PMT 26 by means of graded shield 46 having three layers of metal with very dirr~,lcll- atomic numbers. Typically, such a graded shield includes about 0.2" of Pb, 0.15" of Sn and about 0.1" of Cu.
PMT bases 30 are preferably fabricated from selected materials with low radioactive background, e.g., using pure copper or al~ -i----.-- for the ~7u~po~ g frame. The use of resistors and capacitors selectç(l for low radioactive bac~l~,ulld is disclosed, as well as the use of In-free solder, e.g., made of pure Sn or Sn/Pb alloy. All passive and active elements of the PMT base 30 are se!ected to have a very low tenl~el~Lule drift, and active co---pç~ lit)n trr~ niqlles to eli--.ii-~lc tcl~erdlulc dependent gain drift are disclosed.
Both an OR and an AND mode are used for data acquisition and analysis, with a mnltir~nn~l DSO 52 for on-line background rejection. Triangular shaping and software rejection of fast pulses due to signals ind~lce~ by cosmic rays in the PMT's 26 is employed.
A pulse rise time of about 0.75 microseconds and a slow fall time of about 5-10 microseconds are plcr~ ,d.
Self-diagnostic and self-calibration is used for reliably m~tcning the count rates in the OR and AND modes, more specifically, on-line baseline restoration and pile-up rejection tec~niqlles. In the coinr~ nt mode, it is crucial to use the DSO 58 for m~tclling the shape and temporal coinri(lenre of pulses from the two detector modules. There is a trade-off between the need to estim~te the pulse coinrirl~nre to within better than 100 nsec and the need for a triangularly shaped, long duration pulse permitting rejection of dark cullcll~s from the PMT's. On-line software based pulse fitting procedures however overcome these conflicting requirements.
Pulse shape analysis: Data arqui~ition in a twin scintillator system is based onamplifying and shaping the signals from the PMT of each detector and building a combined energy spectrum for subsequent analysis. The counts in an a~ pri~L~ energy region of interest (ROI) for the desired isotope are then integrated to determine the count rate. For a CA 0223629~ 1998-04-29 sandwich detector con~ g of 2" ~ m~ter NaI(Tl) or CaF2(Eu) scintillators, the detection eff1ciency is typically about 50% when only events in the single-photon energy ROI are counted. For Il25, coulllillg both 30 and 60 keV events increases the detection efficiency to about 70%, but effectively doubles the integration energy range and therefore the background.
Conversion of the count rate (counts per minute, or cpm) into the actual activity in the sample (decays per minute, or dpm) requires knowledge of the detection efficiency (DE) of the counter. For Il25, the DE can be del~ ,ined from the spectrum itself using the known Eldridge forrnula. The DE is d~lellllil~ed for each detector sep.~a~ly, which allows improvements in the calibration and accuracy, and testing the system inl. glily and col-~ g 10 for the sample pl~cem~nt by co,-~ ;"g the two estim~tes of the actual activity of the calibration sample. The ~e~ ~ll of coinri~l~nt events can be used to enh~nre DE evaluation and for diagnostic purposes.
The predominant colllpol~lll of the non-radioactive background in low-energy gamma/X-ray detectors is due to dark pulses in the PMT's. In the Il25 ROI, these typically 15 produce a few cpm for a two-PMT system. However, the shape of these pulses is dirr. .c;nl than those produced by scintillation in the detectors, making pulse-shape based disc~ ;on possible. Pulse shape(s) for each event are acquired using a PC-based dual input plug-in DSO
card 58 and fast pulse shape analysis is performed. This allows rejection of PMT dark pulses as well as other elecLl--,n~gn~tir and vibrational artifacts. After pulse-shape based rejection, 20 the background in the system is almost flat for energies in the 15-100 keV range and is rem~rk~hly stable, independent of the activities in the vicinity of the detector.
Detectors: Flat 2" ~li ",-o~l detectors (1 mm thick NaI(Tl) or 1.5 mm thick CaF2(Eu)) are pref. lled. Smaller scintillators decrease the DE of the system while for larger crystals the signal to background ratio f~ kS. The scintillators are coupled through quartz windows 25 3-5 mm thick to 2" ~ m~ter high energy resolution PMT's which are selected for low background. The PMT signals are read out, amplifled and shaped using ele~;llul,,cs built into the PMT bases 30. To reduce the flux of background photons from the bases to thescintillators the bases are isolated from the PMT's with 5 mm of lead and 1 mm of copper plating with holes for the PMT pins.
Shields and holders: The detector assemblies are placed in graded lead+tin+copper shields (2" lead, 1 mm tin, 1 mm copper). The detectors are placed face-to-face half an inch from one another and a sample holder/crosstalk eli",i"i1lol is placed between them. This is a 1 mm thick copper sheet mounted in a lead frame. Openings are left in the copper sheet for CA 0223629~ 1998-04-29 the samples. The shape of the oyenillg is ~el~ d by the format of the s~mrlec to be used.
A delrin guide with copper outer jacket ensures that all samples are centered in the detector system. When the sample format and sample holder are changed, the system can be ~tom~tir~lly recalibrated using the Eldridge for nula.
Data ~cquicition - hdldwal~e: The data arqllicition haldWdle iS preferably mounted inside the de~ te~l PC controlling the MPD. With lef~ ,nce to Figure 7, the data acqllicitiQn electronics in an exemplary embodiment include triggering circuit 56, amplification/att~ml~tion modules for each detector, digital timer/coull~ 57, and a dual channel 20 MHz digital storage oscilloscope (DSO) 58 which is used both as a 2-input mlll~ l",Pl analyzer and a pulse shape analyzer.
The triggering circuit 56 produces â rectangular trigger pulse whenever a pulse e.x~ee~ing a preset threshold amplitude is legi~ d in either detector 26. If pulses are registered cimnlt~nPously in both detectors a higher amplitude trigger pulse is produced. It is thus possible to count se~alately coinrident and non-coincident events. The trigger pulse is sent to the external trigger input of the DSO. The Ll;gg~ g circuit may be an integral card as ~eccribed below.
The amplificationlâttenll~tion modules adjust the amplitudes of the pulses so that the energy region of interest is within the 0-1 Volt window of the DSO and particles of the same energy produce pulses of the same amplitude in both ch~nnPlc.
The first of the timer/coullL~ls 57 is used as a precise ~cqui.cition time timer (counting the 2.5 kHz refel~ ce pulses). The second timer/counter counts all trigger pulses produced by the triggering circuit, while the third countc only triggers associated with coincident events.
The data acquired from these counters are used to directly evaluate losses due to acquisition system dead time and thus enable the system to correctly count high activity sources.
The DSO 58 (preferably a c~ el.;ial CSLite PC add-on card m~nllf~rtnred by Gage Inc.) is capable of cimlllt~nPous sampling of two input channels with 8-bit accuracy and up to 20 MHz sampling rate and has an additional external trigger input. The data are stored in on-board memory and can be k~r.,lled to the host PC RAM by standard memory-to-memory k~irer via the PC bus. The dead time is strictly non-ext~n-l~ble and by means of the counters described above the count Mtes can be corrected for dead time losses. The DSO 58 is rearmed and initi~li7-ocl after each acquired and processed event.
- The pulse traces are k~l~ç~llcd from the DSO 58 to the host PC 59 memorv and are analyzed for amplitude and shape by software.

CA 0223629~ 1998-04-29 Initially the DSO 58 is set up to continuously chart the input voltages in the two çh~nnPlc and wait for a triggering pulse in the trigger input. When a trigger pulse is registered, the DSO 58 is allowed to capture a prede~P....ii.P~l number of post-trigger points and is then stopped. The relevant portion of the traces (typically 20 pre-trigger and 108 post-trigger points at 20 Mhz sampling) is transferred to the host PC 59 memory for analysis. The rcl procedure takes less than 200 microseconds per trace for a 486 DX66 co~ el for example.
Signal Conditioning/Processing Card (SCPC) 56: To enable the MPD detector software to process signals from two detectors, the trigger cil~;uilly is used to:
1) gel~ te a trigger signal whenever a pulse is produced by either PMT;
2) match the analog gains in the two channels, and 3) adjust delays between the signals and the trigger pulse.
This ha~-lwale (see Figure 7) is preferably implemPntPcl as a signal conditioning/processing card (SCPC) 56 placed inside the colll~ er. The SCPC card 56 features a~ ct~ble gain and trigger levels for each channel and produces a TTL trigger pulse.
The SCPC card receives as inputs the signals directly from the PMT bases 30. Theconditioned signals and the trigger pulse are passed directly to the inputs of the DSO card 58.
The two-channel SCPC 56 is ~lecignP-(l for:
* Production of an analog trigger which starts the DSO 58 both in coincidence mode and the mode of ~ on of input signals.
* Production of a TTL trigger which starts the counter 57 in both modes.
* Blocking the trigger if any input signal has an amplitude greater than some fixed level.
* Analog delay of the input signal.
* AdjllstmPnt of the DSO input signal level.
The SCPC 56 has the following pala~ ,tel~:
* Input signal (positive pulses): 0.1 - 8 V amplitude; 0.1 - 100 ms pulse duration * Lower level adjnctmPnt 0.2 - 5 V continuous.
* Upper level adjllstmPnt 0.2 - 5 V continuous.
* Output attPnll~tion range: --3 to--60 db.
* Digital delay range: 0.1 - 10 ms.
- * Analog delay duration: (1 + 0.01) ms.
* Output signal (positive pulses) CA 0223629~ 1998-04-29 analog signal: 0.1 - 1 V amplitude; 0.1 - 10 ms pulse duration TTL level: amplitude 2 4.5 V; duration 4 - 5 ms.
analog trigger: amplitude 0.5 V for "OR" mode, 1 V for "AND" mode duration 4 -5 ms.
* Telllpelalulc stability threshold level < 0.05 %/~C, transitioncoefficient < 0.01%/~C.
A method in accordance with the present invention will now be described with 10 ~crerellce to the exemplary flow chart contained in Figures 8a to 8c. It would be ap~dlc from the following description of this exemplary method, that the method could be implrmrnted in a variety of processing ellvhollllRnts with a variety of software platforms, and such are considered to be within the scope of the invention.
The on-line pulse rejection soflw~lc rejects pulses whose shape is not colllpdlible with 15 pulses produced by particles stopped in the scintillation detector. Most of the rejected background is due to noise in the PMT, ele~;llo".~gn~tir hll~r~ ce and vibrational noise.
For a given detector, the shape of the pulses caused by r~lioactive events does not change with time and does not depend on the amplitude of the pulse within the linear range of amplification. Thus, one approach to pulse shape rejection is to normalize the registered 20 pulses to a common amplitude and to compare the shape of the norm~ e~l pulse to a "standard" shape using, e.g., a chi-square test. This however, involves lnas~ive floating-point collll,uktlions which are much too time con~nming for effective implementation on ~;ull~ ly available low-cost microprocessors.
Allell~liv~ly, the pulse shape rejection can be pelrolllled by del~llllil~illg a number of 25 parameters related to the pulse shape which can be computed using predomin~ntly or solely hlt,rgel alill----rti~.
Data processing: After initial steps of setting the DSO for aquisition and waiting for a trigger, analysis begins with Colll~u~aliOn of the baseline and the pulse amplitudes in each detector (Steps S3 to S14). From this it is known whether the event occurred in detector A, 30 detector B or both. The pulse amplitudes are adjusted for the current baselines, and if the latter are unacceptably distorted the event is rejected. Then, a number of pulse shape - parameters are evaluated and compared with the ranges of acceptable values established by the CA 0223629~ 1998-04-29 software at system set-up. These include, for example, pulse widths at 1/4, 1/2 and 3/4 of peak pulse height (Figures 8b and 8c). The parameters may include:
pulse amplitude;
rise time fall time total pulse width;
shape of the rising part of the pulse;
the shape of the falling part of the pulse;
the delay bclwcen the pulses from the two detectors;
pulse multiplicity;
the pre-pulse trace; and coll~alalol of the pulse shape with a standard high energy photon pulse shape using a least squares t~c~niq~nP.
Fast integer-based routines for these cc,l,l~u~lions run very effl~ nt1y on Intel 15 processors. For coin~ pnt events (Figure 8b), an additional rise-time ~lignmPnt check is p~.rulllled. After pulse-shape, pulse-height and coincidence/anticoincidence analysis, an unrejected event is added to the a~ lialc spectrum (spectrum of detector A, ~ecllulll of detector B, or 2D ~peCIlUlll of coincident events).
The DSO 58 acquires 128 points with 8-bit resolution for each input charmel for each 20 pulse. The time window spans the whole width of the shaped pulse (S microseconds) and approximately 1 microsecond before the rise of the pulse. Digital procçssing of the pulse begins with c~lcnl~ting the average of the first 8 points S4, which gives the current baseline.
If the computed baseline differs ~ignifir~ntly from 0, the event is rejected (steps S5 to S7) as distorted by pile-up. Next 120 points of the trace are sc~nnPd (using the fast LODS instruction 25 of the 80X86 processors) for the maximum value. If overflow is det~cted, the event is rejected (steps S5 to S7). Otherwise, the amplitude is computed as the dirr~.cnce between the peak value and the current baseline (steps S8 to S10). The program also stores the time offset of the peak. A fast (LODS-based) scan is then done forward and backwards from the peak to detçrmin~- the time offsets of the crossing-points for the levels, for example, of 1/4, I/2 and 30 3/4 of the amplitude (steps S15 to S20, and S26 to S28). These data are sufficient for p~lrlulllillg pulse shape rejection tests.

CA 0223629~ 1998-04-29 After an event has been processed, the DSO 58 is reset for acq~ ition of the next event. Any events oc~ g during the proces.~ing time are lost (dead time). When the spectral data are processed the computed count rates are adjusted for these losses.
The acquisition can be preset to collect data either for a given interval of time or until 5 a certain number of counts within a se!~cted ROI have been acquired. The acquisition can also be te~ tecl by the user at any time.
Calibration routine: The aulolllaliC device calibration/ROI setting routine is as follows.
The user can request the software to p. .rOllll this procedure at any time. No calibrated sample is needed, but a reasonably high activity (optimally 50,000 to 200,000 dpm) I'25 sample is 10 required. After placing such a source into the detector the program acquires 100,000 events with all pulse-height and pulse-shape rejections enabled. Once the acql~i~ition is fini.~h~d the ~,~e~llulll is analyzed to del~ ...inP the region of interest (ROI) for a single photon (27-35 keV) peak. The count rate (cpm) within this ROI is riet~ ~ "~ d. If the actual activity (dpm) in the sample is known, the detection efficiency could be established from the ratio of cpm/dpm.
To estim~tr the absolute activity of the calibration sample, the program now starts a second round of data arqlli~ition with pulse-shape rejection disabled (as the calibration sample activity is high, there is no need to reject the background, and no real events are discarded). Spectra for both detectors (A and B) are built until 200,000 events are acquired.
Subseql~Prltly the spectrum of each detector is analyzed to estim~te the count rates in the 1-photon and 2-photon peaks (the coincident spectrum is also used to enh~nre separation of these peaks). Then the Eldridge formula is used to estim~te the detection efficiencies of the detectors and the decay rate for the calibration sample. The data are chrrl~od for con.~ ei-ry by comparing the estim~tes from detector A and detector B, and the average of the two is accepted as the absolute activity, from which the detection efficiency is estim~trd.
The DE is normally in the range of 50-60% for 2" systems. Only the 1-photon peak is used for counting, which in the case of a sandwich detector with electronic noise rejection decreases the detection efficiency only by about 15% while ~ hing the background about 2 to 3-fold.
After the ~lltoc~libration/ROI setting is complete, the system is ready to count samples of t_e same geometry as the last calibration sample used. Acquisition is pelrolllled either for a preset time or until a preset number of counts (dele----i~ -g the st~ti~tir~l uncertainty of - counting) have been acquired in the ROI. The program then esfim~tes the actual decay rate in the sample using the previously computed DE. The sample counting data are then typically CA 0223629~ 1998-04-29 stored to disk in an ASCII data file which can be transferred to a ~l~t~b~ce or spre~lsh~et program for analysis.
The software sets the ~rquicitionlrejection parameters for each newly assembled detector system. This (lete~ Ps the optimal trigger levels and pulse shape rejection S parameters for the system and creates internal data files to store these parameters. These parameters generally do not have to be re(l~terrnin~-~l during the lifetime of the system unless a major component (e.g., a PMT/base or DSO card) is replaced.
Data pl.,se.lLalion: The system program can be pre-set by the user to count more than one sample or to count the same sample repeatedly a requested number of times. If a batch 10 of samples is counted, the program plO~ the user to change the sample after each measurement, and all data are stored in the same ASCII file. The file contains a header with a date/time stamp and system settings, as well as an extended description of the sample(s) entered by the user using a GUI edit window before the counting begins, for e~ lc.
The program may also be provided with a simple data file bl~wsel which allows 15 viewing and analysis of data from single samples and batches of c~mples. The user can select a data file to be viewed through a system of menus, see the counting ullce.L~ y for each sample, plot the count and decay rates vs. sample number and print out the data with st~ti.ctir~l uncertainties. A more e~L~nsive analysis and merging of data can be performed using a commercial spre~lch~et plVgl~llll, for example.
The software is preferably coded in Borland Pascal and Assembly language to speed up the pulse processing and operates under DOS using a Windows-like GUI shell.
Alternatively, software under MS-Windows can use the recently released Delphi software development system (Borland Tntrrn~tional), which uses the extended Borland Pascal language.
Those skilled in the art could use other programing l~n~ges and stay within the spirit of the 25 invention.
Self-calibration and self-diagnostics: MPD devices according to the invention include self-diagnostics and self-calibration using the re~lllntl~nry in spectral h~lllldLion characteristic for EC sources, i.e., the one- and two-photon peaks in single-detector spectra and the 2D
spectrum of coincident events (the scatter plot of energy in detector A vs. energy in detector 30 B). Additional benefits stem from the MPD using two sepaldl~ pulse counting subsy~L~llls, namely the DSO 58 and the pulse counters 57, which allows molliLolhlg of disclepdllcies in - the count rates bl Iweell the two detector ~ub~y~L~;llls and to detect possible hardware failures.
The MPD software uses data acquired from a source of known isotope to check the operation CA 0223629~ 1998-04-29 WO 97/16746 PCTrUS96/16968 of the detectors, PMT's, HVPS and the readout elecllul~ics. Possible co~ ion of the MPD is ~letected by m~ lring and analyzing background spectra. The software autom~tirally coln~ules the detection efficiency in both the coincident and non-coincident modes for a given EC isotope using a calibration sample of that isotope. Diagnostic lûulilles track any drift in 5 the gains and thresholds in the device and c~lr~ te any required adj~
After tuning, the MPD has ex~ell~nt long term stability; short time (less than 1 week) drifts are less than 1% in the I'25 peak. 1,200 inllPpentlPnt measurements pelrollned with MPD show reproducability is much better than the st~ti~tir~l uncertainty of these measurements. MPD devices permit better than 1% mea~.ule~ precision for sources in the 0.1-1,000 picoCurie range. Over six months of operation the drifts were smaller than 3 % .

For measurements at the zeptomole level the MPD device often runs overnight7 so that diurnal variability is illlpolL~I. A 5~C tellll~el~lule change leads to a noticeable, about 1-2%
drift in scintill~tor/PMT assembly pelrollllallce. To colllpensdle for this effect the telllpel~lùle of the crystal can be measured and used by the software to correct pulse amplitude.
Asymmetric detector geometry will now be ~ cllssecl. The single sample prefelredimplem~nt~tion discloses a fully symmetric sandwich configuration of the MPD. This geometry is ~lllul,liate for the symmetry of decay of CGX isotopes; there is no correlation in the emission direction b~l~een the coincident photons. For I'25 the two photons are nearly the same energy, which allows optimal pelfollllance using symmetric detectors. The use of any other geometry for ~,y~ ;r decay is thel~fol~ inefficient. There are, however, applications which justify the use of asymmetric configurations of the geometry for best p~lrollllance. The various types of assymetry which can be used according to this aspect of the invention advantageously include but may not be limited to the following:
* the two detectors are made from dirr~,lell~ materials, e.g., two dirr~relll types of scintillator can be used, or one scintillator and one semicon~iUcting detector may be used;
* the two detectors are of dirf~ lll size, typically one very small to tlimini~h background and one large to increase the detection efficiency;
* the two detectors serve dirrelellt functions, e.g., one is spatially resolving and other is a non spatially resolving detector which serves as a triggering device. Typically, the spatially resolved detector is partially obscured by an applopliate pattern of coded apelLules while the second detector is fully exposed to the source.

CA 0223629~ 1998-04-29 Optimal configurations use a combination of el~mPrltc. In one impl~ ;1linn according to the invention one detector includes a NaI(Tl) crystal coupled to spatially resolving PMT. However, the triggering detector is a CaF2(Eu) scintillator coupled to a dirÇel~nl type of PMT from another producer.
S For a spatially resolving MPD detector (herein refeITed to as SR-MPD), the system pclrolll~ce depends on optimi7.ing all parts of the system, e.g., the type of scintillator, scintill~tor rli~mPter~ scintillator thirl~nPss, thirl~nPss of the optical window, and the type and geometry of the coded apellule mask.
A high resolution, spatially resolving MPD is now described. The single sample MPD
10 permits considerable bacLgloulld reduction when using a single sample labeled with CGX
isotopes, especially I'25. However, for many applications, one needs good spatial resolution, e.g., when mP~cllring radiolabeled di~llibulions obtained in separation processes (electrophoretic gels, dot blots, thin layer and paper chlollldtography), and in anatomic studies of tissue pl~p~u~tes.
Very popular 2D radiolabeled di~llibulions include products of separation by electrophoresis, thin layer chromatography (TLC) or high perfollllance liquid chl.,ll~lography (HPLC) which can be subsequently filtered upon an a~roplidte mP~inm. These may be referred to as chl~,llla~grams. They are usually self-supporting, i.e., the separation product is deposited or trapped upon the surface of a thin, mech~nir~lly stiff plate. The most popular class of 2D radiolabeled distributions are electrophoretic separation products trapped inside gels. Typically, the gels can be cured or dried to hlllJiove their mPrh~ir~l pl~.,.~ies. In this and many other many cases, however, it is more practical to sandwich the biome-lic~l sample between two thin films of material. Finally, electrophoretic separation products are often r~ ,d or blotted upon elastic membranes, either nitrocellulose or plastic. These products are called blots.
For all these applications, MPD detectors can be implemPntP~l to replace cl~c~ir~l autoradiography and phosphor imagers. In all these applications the biomP-lic~l sample is essentially a 2D object, thir~nPsc being much smaller than the two other dimensions. Often, it is supported on one side by an apl,lopliale thin membrane. In the majority of applications, the thin support may be produced from a material with low atomic number, e.g., plastic. Such a thin mPrh~nir~l support can be implemented so that it has relatively small absorbtion of X-rays, say less than 5 % .

CA 0223629~ 1998-04-29 W O 97/16746 PCT~US96/16968 Three colllpelillg re.luilel.lc"L~ on the physical plese~ l ion of radiolabeled 2D
distributions are:
1) samples should be flat and easy to handle mrçh~nir~lly;
2) samples should be herm~tir~lly sealed to elimin~te detector co-~ ion by direct S contact or by degaccing of aerosols;
3) the X-ray absorbtion in the col~ing material should be minimi7etl.
These three requirements are due to the unprecedentPcl techni~l performance, especially sel~iliviLy, of SR-MPD detectors. First, the sample should be as flat as possible, because a change of ~lict~nre from the sample to the detector surface leads to artifacts in the measured activity. Conventional detectors are typically calibrated to within +5% whereas the MPD
achieves +1% calibration and reproducibility. Also, when samples are not flat there is considerable loss of spatial resolution. The unprececlented sensitivity of the MPD means that even the .cm~ st co--l~.--in-~iQn is fatal for measur~ ent integrity. Often, not only the biomaterial but also the supporting m~tPri~l is co--~.--in~l~d. This is the case for blots because the blotting is pelrol,lled using a special buffer liquid, and a small fraction of radiolabeled electrophoretic product is transported by diffusion to the reverse side of the membrane.
Similarly, the reverse side of TLC plates is often slightly co-,~ ed. Finally, when c~elaling with sub-picoCurie samples, any additional absorbtion is a problem. Cc"~r--r~ nt methods using very thin layers of low atomic number materials are therefore p~crt;llcd.
Three ~lcfellcd methods of enr~ps~ ting 2D radiolabeled blots are as follows. For self-supporting samples, e.g., chlolndl~graphic plates or blots, the easiest and most practical method is to spray them with a liquid lacquer. Acrylic sprays, silicon sprays and GE electrical vernix have been used s~rces.cfully. It is important to check the spray material for radioactive co--~ ion; and to use only sprays which have coincident activity lower than 0.1 picoCurie/cc. Another practical method is to use thin adhesive tape made of either plastic or very thin ( < 0.1 mm) ~ll-...i..l-.-. tape. Thin pouches may be used, preferably pouches made of thin films of beryllium. However, for most applications pouches or bags made of plastic or very thin ~l.. i.. l.. are a~l~qll~te. For non self-supporting samples, l~min~tion is a very practical method of providing them mrch~nir~l plopt;llies and radiorhrmir~l purity required 30 by the SR-MPD instrumentation. As l~min~tion is typically achieved by rolling the foil-sample-foil-sandwich bclween two hot surfaces, care should be taken that the l~min~tion - m~rhinr itself is not cont~min~ted. Also, care should be taken that the l~min~tion foils are not co~ ted .

CA 0223629~ 1998-04-29 A sc~ g MPD device is now described. In most symmetric MPD systems, spatial resolution is comparable to the crystal ~ mPter. For low energy X-ray e.l.ill~l~, e.g., I'25, the spatial resolution can be drastically improved by inclusion of a "pin hole" or "slit"
~ellulc. Then, 2D isotopic distributions can be m~ch~nir~lly displaced between two S detectors, one of which has the above said ap~ ul~. This implementation is referred to as the MPD-Scanner.
It is (liffirlllt to obtain sub-millim~ter resolution without losing the se~i~ivily required for i~ cLillg 2D radiolabeled biomolecula distribution, e.g., DNA disl ibulions at sub-attomole levels. In conventional imagers for these applications, the principle of proximity 10 im~ging is implem~ntçd For example, in spatially resolving detectors, for beta e.,-il~.~
"system resolution" depends upon the energy of the used emitter; it is con~ rra~ly worse for higher energy sources due to an increase in the range of the beta particles. For example, S35-labeled DNA bands are sharper than P32-labeled DNA bands when imaged with film or a phosphor imager, even though they are i lt~,rogaling the same DNA band. Thus the "system 15 resolution" of static systems depends not only on the intrinsic resolution, but is considerably broaden~d by the emission pellulll~ld.
The MPD-Sc~nn~r according to the present invention permits dynamic data acqlli~ition with excellent S/B. It allows the system resolution to be directly proportional to the precision of the s~ lu.e movement upon the surface of the blot or other 2D sample format. Typically 20 the collimator (slit) width is of the same order of m~gni~ as the characteristic physical band width of the sample. A stationary collimator measures the total signal coming into the entire slit area, e.g., the signal from 4 mm2 of the sample for a 1 mm x 4 mm slit. The situation is dirÇe.el.l when time resolving detectors are used. As the collimator moves to a new position across the blot, one measures a relative increase or decrease of activity in the area of the blot 25 that is opened to detector by the movement of the leading edge of the slit. On the other hand, some part of the blot is no longer exposed because it is now covered by the trailing edge of the slit. Thus, knowledge of the time of signal arrival permits dirr~re..lial deconvolution of the blot activity and sub-millimrtric spatial resolution when im~ging the blots. The "system"
spatial resolution is directly pl~o-lional to the accuracy of the movement of the sc~ulel, the 30 MPD-Scanner can provide about 100-micron accuracy.
When using a moving slit with width x, the spatial resolution dx is of the order of (x/A) where A = min{S/B; sqrt(N)}, where S/B is the signal to background ratio and Sqrt(N) is the st~ti~tir~l ul~ellaillly of the measurement when N photons are detçctçd for a given slit CA 0223629~ 1998-04-29 position. Typically, S/B > > sqrt(N) > > 10 and dx is approximately 0.1 mm. For 30 keV
photons, typical for Il25, about 200 microlllcl~l~ thick t~lngcten foil stops over 90% of the photons. Thus the "edge effect" will limit resolution to about 100 micrometers. This limit can be as low as 20 microlllel~ for lower energy EC isotopes. There is an analogy between S the improved resolution of the MPD-Scanner and the confocal microscope; the spatial resolution is better than but proportional to the a~ ulc dim~n~cion There is an appdlclll limitation to such a system; the count-rate is drastically reduced.
The ~p~illlulll 11im~ncions and material of the ap~lluic are dependent on the application and the activity of the 2D distribution. Plcf~.led are lead films with thirl~n~ss from 0.2 to 1 mm.
10 Typically, for sc~ TLC plates and agarose gels, an apellule in the form of a slit with an ope' ~ width of 2 mm and length of 2 cm is plef~.lcd. For higher resolution acrylamide gels and sequen~ing blots in general, one can use slits with 1 mm width and a few mm length.
Using the MPD-Scanner, the spatial resolution is much better than the a~ ulc ~lim~ncions; typically 0.2 mm resolution. However, this case involves reconstruction sorl~alc 15 in which for each step of the m~chqni~ql mover, the optimal count-rate is cql~ulqt~d. This optimal count rate depends on the signal-background ratio at the given place of the 2D
distribution.
Thus, the plefill~d scan routine is iterative. First, a low precision, constant step stqti~tirqlly limited map of the 2D isotopic distribution is obtained. Arlc~ lds, an o~ cd 20 scan is pelrollllcd in which the acquisition time at each spot is cq-lrulqtecl to oplillli;~e the total scan time under the constraint of minimql spatial resolution and stqti~tir~ql ul~hl~y. The operator may choose some part of the 2D sample to be scqnn~d with higher precision.
The counting rate in the MPD-Scanner depends on the ~lim~n~ions of the crystal. An optimal configuration is one in which two crystals have dirr,l~nl rliqmPter. The detector on 25 which the apc,lule is placed is smaller, typically 0.75 or 1 inch in rliqm~er. The second crystal is much larger, typically 3 or 4 inches in diqm~ot~r. The MPD-Sc,q-nn-or should be a~yllllllc~ic not only in rlim~n~ions but in the use of dirrelcll~ scintillators. For example, signal/background can be improved by about a factor of two when the smaller crystal is NaI(Tl) and larger is CaF2(Eu).
Software for the MPD-Scanner: The two detectors used in the MPD-Scanner according to the invention are the small ~ qmpter (0.5-1 inch) ~lhllaly detector which det~rmintos the - spatial resolution, and the secondary detector used for coincidence/anticoincidence analysis.
The surface of the primary detector is covered with an absorbing mask (of Pb or Cu) with an CA 0223629~ 1998-04-29 WO 97tl6746 PCTAJS96/16968 ap~,.Lulc (usually in the shape of a l~;Li~*ular slit). When scan~ .g is pelrollned, the co~ tel-controlled mover moves the sample in front of the slit with a step which can be equal to or smaller than the slit width. At each step a mea~u~ L is performed, and subseq~le~tly the profile is reconstructed.
S The aCq~ iQn logic is as follows. The triggering is pelrul"led from the signal in the plilll~y detector. Each pulse from the ~lhll~y detector is analyzed for the shape (for background rejection) and amplitude. The trace recorded simllltenPously from the secondary detector is also analyzed, and coincidence/anticoincidence analysis is ~elÇoll.~ed. The energy spectra of all non-rejected events in the plullaly detector and of coinrident events are built 10 during the meas,lre nent. Before the sample is moved to the next position, the counts in ROIs set for dirr~lenL isotopes are integrated and the data appended to the scan data disk file.
The user interfare for the MPD-Scanner inrludes the scan definition and the dataanalysis modules. The scan definition module permits the user to preset the geometrical palalllettl~ for multiple scans: the starting points, the scan lengths and steps, as well as the 15 measurement times per step (which can be dirr~.e.ll for dirr.,le.ll scans), and the user descriptions for each scan. The data analysis module permits viewing the scan profiles in dirr~lell- modes (a profile graph or a sim~ ted autoradiogram). This module also permits analysis of profiles, as well as sending the data to a spre~chret for customized analysis.
Very high spatial resolution sc~nnin~ MPD is now Ai~rll~sed In many applications, 20 a few microns spatial resolution is required. An MPD with such spatial resolution permits considerable improvement in a plurality of biomPAir~l studies, e.g., ~n~tomir~l and cellular studies. The spatial resolution of MPD-Scanners is generally about 100 microns. The spatial resolution of all detectors is limited by the following effects:
1) thir~nrs5 of the 2D distribution of the radioisotope.
25 2) the limited ~lo~ g power of the detectors, leading to parallax errors in the crystal;
3) limited stopping power in the apellul~e, leading to a non-delta llallsÇel function;
4) positioning errors in the m~rh~nical displacement system.
The first error source can be elimin~t~d using devices analogous to those used in conventional optical and electron microscopy. The second source of error is a fimA~mrntal 30 limitation leading to positioning uncertainty in all detectors using proximity im~ging. This is the main limitation upon the spatial resolution of autoradiography or phosphor imagers. As - mentioned above, in the case of beta sources, the range of the particle limits the resolution of classical detectors for all sources but tritium. Obviously, there is a trivial solution, which is CA 0223629~ 1998-04-29 W O97/16746 PCTrUS96/16968 to make the scintillator as thin as the required spatial resolution, but this leads to very low detection efficiency.
Use of im~ing with apc.lu~es partially removes the problem of the range of particles.
This solution is especially attractive in the case of soft X-rays whe~eil1 the stopping power of 5 gold or pl~timlm film is about 30 times higher than of NaI(Tl) and up to 100 higher than for CaF2(Eu). However, even in the case of heavy metal films, e.g., gold, pl~tinnm, tlmgst~n or lead, a large fraction of the 27 keV photons pass through a 100 mitrons thick foil. Software which permits creation of images for partially ll~nsl~clll films i~ roves the spatial resolution of MPD-sca~ only down to about 50 microns for Il25.
For further el~hA,ll~e~,.f ~~l a l.rer~llcd solution is to (limini~h the energy of the photons used in the image creation. These are found in EC el~ with lower atomic ~lulllbel, e.g., Fe5fi or Cr5'. In this case, about 6 keV photons are present, which using gold film can be imaged with a precision of better than 10 microns. Another alternative is the use of Il25 and Il23. In this case, not only the L-edge characteristic photon~s (27 and 31 keV) are present, but also 4 keV K-edge X-rays. Finally, one can use Auger electrons emitted by higher energy CGX sources.
In the case of very soft X-rays, say < 15 keV, sçintill~tors are not the best deleclo- ~.
One can use a hybrid MPD-Scanner where a small silicon detector is covered by a high density film with apc.lu,c. The second detector detects only higher energy photons, e.g., 27 and 31 keV photons in the case of Il25. Thus, it can be a relatively large, say 2-3 inch ~ m~ter, scintillator, e.g., CaF2(Eu).
Al~e,l~lively, a gas dclector can be used for detection of very low energy X-rays. A
windowless gas detector can be used using precsnri7~d gas flowing through the film aperture.
This film itself is used as an electrode of the gas detector amplifying system.
A fourth source of errors in the MPD-Scanner is the precision of the m~ch~nir~l displ~trm~nt system. Pie7.oelectric m~ch~nir~l displ~rem~nt generators exist with sub-micron resolution. Additionally, the problem of the precision of m~ch~nic~l parts of the MPD-Scanner is facilitated in those applications where only relative, rather than absolute indexing, is required.
An exemplary Spatially Resolving MPD (SR-MPD) is now described. The Scintillator-based Spatially Resolving SR-MPD detectors feature low cost, excellent sel~silivi~y and good - spatial resolution. The SR-MPD con~;ul,c,.lly (~ lPs up to 50 samples of biological material labeled with CGX isotopes, e.g., 1125, with reasonable detection efficiency and CA 0223629~ 1998-04-29 minim~l cross-talk (below 1%). The bac~loulld is about 0.3 count per sample per day. The spatial resolution of the SR-MPD is about 2-3 mm. The SR-MPD can be s~lccec~fully applied to analysis of biologicals presented in the form of dot blots or contained in standard 8x12 well Illicl-~liL~l plates.
S A p~f,lled SR-MPD according to the invention includes t_ree parts: an auxiliary detector, a spatially resolving detector and data acquisition ele.;Llol~ics. The auxiliary detector uses a 3" CaF2 (Eu) scintillator coupled to a low background non spatially resolving PMT.
T_e pG~ lce of the auxiliary detector is ~lJlhl~ ed by selection of a crystal cli~ tçr and thirlrn.oss which provides the best trade-off belw~cll detection efficiency and low bachglvulld.
The methods disclosed for the single sample MPD (selection of low background PMT's, use of thin quartz window, special parl~gin~, telll~lalulc stabili_ation of read-out elecLl~ nics, special design of PMT high Voltage base) are used here. They permit a factor of five decreased background as compared with the colll.lle.~;ially available assemblies. The spatially resolving detector includes a t_in 2-3 " ~ m-oter NaI(Tl) crystal coupled to a spatially resolving PMT (SR-PMT). P~cr~ ,d SR-MPD devices are impl~ ~--e~ l using a 3" (li~mr~r SR-PMT
m~mlf~rtllred by ~ u To count multiple samples with minim~l crosstalk, a lead mask about 1 mm thick is placed on the surface of the SR detector. Coded a~kllule masks made of tll~g.cten, gold or pl~tini~les permit further hl~rov~ lL of spatial resolution. For example, samples arranged as a dot blot are placed next to the ape, IU1~S in the mask. This configuration allows up to half of the single-sample MPD detection efficiency for each of about 50 samples. One of the problems is non-linear SR-PMT response. To m~ximi7,~ the number of samples measured coll.;ul~ellLly the samples can be arranged so that the sample spacing increases towards the outer edge of the SR-PMT. The SR-MPD permits up to 20 times higher throughput than the SS-MPD at about three times the cost.
For I'25, the X-rays are very soft. This permits im~ging when using a parallel hole collimator. The use of SR-PMT's in conjullclion with sc~nning devices permits excellent spatial resolution, down to 0.2 mm. This is a~lequ~te for nearly all molecular biology applications. SR-PMT techniques can be adapted to the exacting standards of ultralow-radioactive background applir~tion~. SR-PMT's have previously been developed for high counting rate applications, typically 100 cpm, whereas according to the invention they are used - at count rates lower than 0.1 cpm. Thus, the radioactive background of the PMT's, vibrational and ele~ gnrtir noise must be rejected and spatial resolution inhomogeneities CA 0223629~ 1998-04-29 and artifacts accounted for. The HA~ U 3" SR-PMT, model R2486, is plefe,l~,d. The electronic readout system of the H~in~ SR-PMT provides the user with four inputs, from which the cool~dilldtes of the ~3etecte(1 events can be c~lc~ t~(l.
The SR-PMT is physically a 3" device which has however, only an approximately 2.5"
5 ~i~mPter active area, i.e., an area in which good spatial resolution can be achieved. The signal drops drastically at about 1.1" from the PMT center which for X-rays leads to drastic artifacts in the measured position of the optical photon cascade in the scintillator. Thus, 2"-2.5" ~ mPt~r scintillators are optimal for the SR-MPD, and good spatial resolution can be obtained by ~l)loplidl~ hal-lwdr~ and by using iterative position calrlll~tion software, 10 described below. For Il25 this means that 49 samples can be well resolved, as co,~,d with the 32 samples which can be resolved using the m~m-f~rtl-rer's elecllollics and the m~n--f~ct -rer's suggested position c~lr~ tion software. From the point of view of background, it is very important that the crystal is ~ignifir~ntly smaller than the SR-PMT
~i~mPter; use of a 2" rli~mPter scintillator leads to three-fold lower bac~gloulld than when 15 using a 3" crystal. Furthermore, the use of a very thin, say 0.5-2 mm thick quartz coupler belweel1 the SR-PMT and the scintillator crystal helps ~limini.ch backgroulld due to beta particles e.,-~ g from the PMT glass about five-fold.
An alternative SR-MPD insLlulllenl is based on the 5" tii~mpter H~ SR-PMT's.
Spatial resolution is only slightly worse than with the H~m~m~t~l 3" SR-PMT. It permits 20 considerably more resolved pixels per detector surface than the 3" SR-PMT. The 5" SR-PMT's are not plc~ d for a low background device. To support the tube against implosion due to atmospheric pres~ur~, the 5" SR-PMT utilizes an approximately 6 mm thick front glass window that is highly cont~min~tPcl with K40, leading to about a factor of four higher radioactive background than with non-spatially resolving 3" PMT's. A larger SR-MPD is 25 preferably based on a 5" SR-PMT made of quartz.
Software for the SR-MPD: The SR-MPD data acquisition software has to process more than two traces for each event. The H~ SR-PMT has four outputs which in thefollowing are referred to as the signal left (SL), signal right (SR), signal top (ST) and signal bottom (SB). These signals are combined in the SR-MPD detector hal.lwale to obtain the totdl 30 signal, TS = SL + SR + ST + SB. Thus, the signal acquisition and procçcsing software analyzes six signals, SL, SR, ST, SB, TS and the signal from the non-spatially resolving trigger electronics (TR). The signals from the spatially resolved detector can have very dirr~lell~ shape than the signals from the auxiliary detector. The signals from NaI(Tl) are CA 0223629~ 1998-04-29 WO 97/16746 PCTrUS96/16968 much faster than from CaF2(Eu). The software uses this dichotomy to better reject the background.
Sum pulse TS and pulse TR from the secondary detector are analyzed for amplitudeand shape to reject electronic and other artifacts, while the four original signals from the SR-PMT are analyzed for amplitude only. The amplitude of the sum pulse is roughly proportional to the particle energy and is analyzed in cle~lirat~d hal-lwalc for compatibility with the source used. However, there is a depentl~n~e of the signal on the photon impact position on the crystal. Thus, after the position of the event is established, the amplitude of the total signal is software analy~d and all events with emlgies outside of the pre-set energy ROI are 10 rejected.
Overall, the dirre~ modes of haldwarc and software event analysis permit rejection of over 95% of the background events. In the case of the non-spatially resolving MPD the main source of background are upsets due to cosmic rays, whereas in the case of the SR-MPD, the main source of background is a combination of elecL~ ~nPtir i-~Ltlr~ ce and dark 15 ~;ullcnL~ in the PMT.
The location of the event on the surface of the detector is cal~lllq-t~ocl from the four outputs of the SR-PMT. The first approximqtion is given by x0 = (SL - SR)/TS and yO=(ST -SB)/TS. However, this approximation is valid only for events with an impact close to the center of crystal, whereas on the edges, there are important position artifacts. Thus, an 20 iLcllaLivc position search routine is plcf. ,lcd, whclcill the real position is established from:
x; = (a[xj l] * SL - b[xj l~*SR)/TS
and yj = (c[yj l] * ST - d[xi l]*SB)/TS .
The calibration functions a(x), b(x), c(y) and d(y) have to be established ell~ilically for each 25 SR-PMT. UnfolL~IaLcly, this function flepçntlC also on the source energy. During data aGq~ ition, a 2D image corresponding to the surface of the detector is built. Following the arqui~ition, the counts in the areas associated with apclLul. s in the mask are illLc~iated to obtain the count rate for each sample, which is then converted to dpm using the calibration data.
The parts of the image corresponding to the apertures and the detection efficiency and background values for each al.,.L,lle are ~çt~-.,-inkd during the calibration of the device by placing samples of known activity into the apertures, acquiring the image, and analyzing it.

CA 0223629~ 1998-04-29 W O 97/16746 PCTnJS96/16968 The software mask construction is pe,r~ led autom~tir~lly by the software using a straighLrcl~lJ peak ~letPction routine.
The MPD Imager is now described. Scintillator-based MPD-Imagers feature low cost, e~rellent sensitivity, high throughput and sub-millimPtri~ spatial resolution. The MPD-5 Imagers can be used for qll~ntit~tion of fractionated biological materials, e.g., chromatographicoutputs and DNA seql~Pnring gels and blots. The spatial resolution of the MPD-Imager can reach or exceed 0.2 mm.
The MPD-Imager permits sub-millimeter resolution and is al,~ro~liate for ql-~ntit~tion of fractionation outputs, e.g., sequencing gels or chromatographic plates. In this device, the 10 SR-MPD is coupled to a high precision 2D mover. High resolution is obtained by using a multi-apclLule pattern; each event is a~si~nPd to a spatial ROI and reconstructed from the knowledge of the position of the mover. The MPD-Imager is much more sensitive and faster than phosphor imagers, which are increasingly used in molecular biology. DNA sequerlring blots were qll~ntifiPd using the MPD-Imager and dot/bar p~tl~ ~ n~ were spatially resolved at 15 few zeptomole level.
A ~lcrcllcd MPD-Imager inrludes the following sub-systems:
~ 2.5" NaI(Tl) scintillator with 3" spatially resolving PMT;
~ 3" CaF2(Eu) scintillator with low radioactive bac~l~lund PMT read-out;
~ a coded a~ellulc/separator/shield ~ubsy~lclll;
20 ~ a 2D mover system with about 100 microns relative movement precision;
~ read-out electronics, inrluAing 3 DSO 58 cards;
~ a data acq~ ition/analysis subsystem (e.g., Pentium 90MHz).
The MPD-Imager permits dynamic arqui~ition of information with excellent S/B. Itallows system resolution to be directly proportional to the precision of the ~cllulc movement 25 upon the surface of the blot. Typically the collimator (slit) width is of the same order of m~nitl]de as the physical width of the DNA band. A stationary collimator measures the total signal coming into the entire slit area, e.g., the signal from 4 mm2 of the blot for a 1 mm x 4 mm slit. The situation is dirÇclclll when time resolving detectors are used. As the collimator moves to a new position across the blot, one measures a relative increase or 30 decrease of activity in the area of the blot that is opened to the detector by the movement of the leading edge of the slit. On the other hand, some part of the blot is no longer exposed - because it is now ~hi~ld~d by the trailing edge of the slit. Thus, knowledge of the time of signal arrival permits dirrclcntial de-convolution of the blot activity and sub-millim~tric spatial CA 0223629~ 1998-04-29 resolution when im~in~ blots. The "system" spatial resolution is directly proportional to the accuracy of the movement of the scal~lel, the MPD-Imager has about 100-micron accuracy.
The intrinsic resolution of the MPD-Imager is now ~ c~lssecl. This parameter can be measured using "radioactive" ink in an HP Inkjet printer to gencl~lc a series of well def~ed S patterns enabling cOlllpdlisOl1 of the MPD-Imager and phosphor imager. A relatively low surface activity of 10 picoCurie per cm~ was used. The test pattern consisted of equally spaced h~ olllal bars of 3 mm, 2 mm and 1 mm, with the bar spacing equal to the bar width.
The pattern and its image were obtained with the MPD Imager and a Molecular Dynamics brand phosphor imager. After a 48 hour exposure, the phosphor imager detected the 3 mm 10 pattern well, although the background is quite high. It only partially resolves the 2 mm pattern, and the 1 mm pattern is lmmP~cllrable. In contrast, the MPD-Imager clearly resolved even the 1 mm pattern with S/B of about 10. The 0.5 mm pattern is also resolved, but requires a longer sc~ g time.
Software of the MPD-Imager: The MPD-Imager combines the functionality of the SR-15 MPD and that of the MPD scanner. The mask on the primary (SR) detector has an array ofdpcl~ules, defining the spatial resolution of the imager. The 2D sample is placed on a c(Jlll~ukl-controlled mover which sequ~Pnti~lly places the sample in the measulcmellL positions.
In each position, tne counting is pclro~llled for a pre-set time (using the same acquisition logic as that for the SR-MPD). The count rates for all al,c~lul~,s are obtained and stored in a disk 20 file.
Following the scan, the image is reconstructed from the coul~ lg data. The resolution-defining apcllulc pattern is such that, to obtain a continuous image, the pixels have to be interleaved in both X and Y tlim~n~ions. The pattern should allow such interleaving, although it does not nPcec.c~rily have to be rectangular. For each available resolution mask, the system 25 has a separate calibration file co--l~;--il~E the software mask defining the mapping of the apellules to the image surface. The mask is constructed autom~ti~lly by the software during calibration.
The user interface allows selecting a lccl~l~ular area to be sc~nnPcl and pre-setting the counting time for each pixel. The data analysis module reconstructs the image from the 30 counting data, permits vi~ li7~tion of the image in false colors, shades of grey, a contour map, or as a 3D surface, as well as storing the image in a number of standard formats for - analysis by other image processing programs.

CA 0223629~ 1998-04-29 A Large MPD-Imager is now ~1i.cc~ssecl. For a large MPD-Imager, the spatially resolving part of the system is çnginPered to enable higher throughput for large 2D
fractionation outputs. Commercial value of MPD-Imagers depends on providing throughput sllffirient to perform overnight analyses of 2D gels and blots at the 10-'9 mole level. To 5 achieve this, one may use a large area, say 12 inch x 8 inch, MPD-Imager.
Three types of large spatially resolving gamma detectors which can be used in a large MPD-Imager are: detectors using one or several SR-PMT's; ~letPctors using a few tens of small ~ m~ter PMT's in an Auger camera configuration; and detectors using mic~ocham1el plates and CCD imagers. Use of a plcr~llcd new scintillator, Yttrium ~h-.,.i.. Perovskite 10 doped with Cerium (YAP(Ce)), permits a large MPD-Imager with very good spatial resolution and exceptionally low bacL~l~.u~d.
The beneficial characl~ lics of a large MPD-Imager are:
1) lowest possible radioactive background, pellllillillg zeptomole sensilivily in a plurality of biological tasks;
15 2) h~~ ion ye~ g high throughput even for the samples with very low levels of radioactivity;
4) low cost, user friendly devices with software facilitating biological tasks.
In one impl~m~nt~tion of a large MPD-Imager according to the invention, a singlecollllJuL~l controls the con-:ul,elll and coordinated operation of four spatially resolving sub-20 units. This is made possible by the relatively low count rates in targeted applications at countrates of between a few tens per second to a few counts per minute per pixel. Pulse shape analysis takes only about 100 microseconds and can be ~limini~h.o~l to about 10 microseconds.
Losses due to pile-up of events in dirÇ~l~lll detectors can be fully accounted for.
The number of detectors in the multi-detector h~Llull~ll~ is limited by the DSO 58 bus 25 throughput, pulse shape analysis time including speed of the controlling microprocessor, the number of available bus slots, and cost considerations. A four detector MPD may be interfaced on a Pentium 120 MHz in full-tower configuration and motherboard with 12 bus slots, for example. Four SR-PMT's can be multiplexed on a single con~ul~l.
Large MPD-Imager based on several SR-PMT's: This exemplary embodiment has four 30 modules:
* auxiliary detector - * spatially resolving detector built of four sub-assemblies;
* m~rh~nir~l mover assembly;

CA 0223629F, 1998-04-29 * data ~cqllicition and procecsin~ unit.
The auxiliary detçctor is based on CaF2(Eu) scintillators coupled to a plurality of selrctçd, low background PMT's. Preferred auxiliary ~içtçctor modules are as follows. First, a large, say 8" x 8" crystal is coupled by means of apploplia~ light guide to a 6" PMT. This 5 is the simplest and lowest cost implementation, but it leads to a rather large device. Second, a single CaF2(Eu) scintillator is coupled to an array of PMT's. For example, a 9"x9"
scintillator can be coupled to an array of nine m~tçhPd 3" PMT's. Third, a 8" x 8" crystal is coupled to the array of 16 m~trh~d 2" PMT's. The outputs of all PMT's are s ~mm~d and pulse height and pulse shape are analyzed by low noise electronics, in~]u-ling DSO 58. Pulses 10 of ~r~liàle amplitude can be used as the trigger for acq~icition and analysis of data from the spatially resolved detector.
It is preferred to use "m~tçhPd" PMT's with serving electronics. When stim~ te~l by a particle of a given energy, the amplitude and shape of the pulses should be çccçnti~lly the same. For example, Iclll~Olal delays should be the same to within 100 nsec, the amplitude the 15 same to within 10% and shape of pulses almost i~ tir~l. To overcome the reqllirmrnt for m~tch-~cl PMT's, the trigger detector can consist of four sub-assemblies, each concictin~ of 4"x4" CaF2(Eu) crystals coupled to a single, 4" PMT.
Preferably, the PMT is square or hexagonal, but cylindrical PMT's can be used with optical coupling using an appl~li~le acrylic waveguide. In the case of a mosaic of four 4"x4"
20 detectors, instead of laborious m~tching of PMT's, much simpler colll~u~el calibration can be used. However, inhomogeneities of scintillation detectors are always largest close to the borders of the scintillator, i. e., the mosaic trigger detector may have somewhat lower detçction efficiency and lower energy resolution, which negatively influences the background.
NaI(Tl) can be used for its good stopping power and the best energy resolution among 25 scintill~tors. An exemplary large area spatially resolving detector module uses a 8"x6" active area to permit im~ging of the full surface of a typical se~lel-ring gel. An exemplary mosaic SR-detector uses four smaller SR-PMT's. The detector assembly includes four spatially resolving detector modules, each consisting of a 4" x 3" NaI(Tl) crystal read out by a ~m~m~tcll square SR-PMT. The square SR-PMT has an active area of 60 x 55 mm, whclci 30 a spatial resolution of 2-3 mm is expected. Square rather than round 3" SR-PMT's are plcfellcd because they permit better coverage of the surface. Both the electronics and the - im~ging ~ro~.,.lies of square and round SR-PMT's are almost identical, and can use basically the same electronics and software.

CA 0223629~ l998-04-29 To economize the slots available in the control c~,---y~ all of the electronics for a single SR-PMT is preferably placed on a single card. The large MPD-Imager uses multiplexing to rl;~ the ~lul~ber of electronics cards to eco,,ullli~ space, facilitates hqn-lling of heat-load problem and ~l;...i.~;~l, the cost of the device.
S Use of a single Pentium 120 Mhz assumes that a single full length card contains the signal conditioning and coincidence cilcuilly for two detectors, i.e., four ch-qnnrlc. Even a 12 slot motherboard lacks enough ISA slots because the DSO 58 featu,~,s two channels per card. A p~crc~lcd DSO card 58 has 4 channels, each 50 MHZ. It is based on CMOS
e1~m~ntc to considerably (l;."i~;.ch heat rliC.ciration. Four chqnn~lc of DSO can then fit on a single full length card with the PCI bus.
Multiplexing electronics is plcfell~d with eight thresholds providing flags for sOflwalc.
For each (let~ctor, there is scy~lc delay and coinri~lenre Cilcuilly, pC~ l;ug both non-coin~idrnt and coinri~lent counting. A set of 40 flags (outputs from 8 thresholds for each SR-PMT, four coincidence signals and four thresholds for sum signals) are then ~ r~,~led to the colllyu~l permitting on-line event reconstruction. The signals from all SR-PMT's are sllmmp~
and the outputs are input into the 3 DSO's. This permits pulse shape analysis using the present software. With the availability of a four channel DSO, only three cards are ~-Pces~y to process the information from four SR-PMT's. By using polarity encoding to process the data from two SR-PMT's, only two 4-charmel DSO cards are npces~y to process the 20 ilrollalion about the pulse amplitude in the SR-PMT's (sum of all 4 outputs) and non-spatially resolving PMT.
To manage thermal load, a few fan cards may be placed bclwcen the DSO and SCPC
cards. The çh~llenge of electrom~gnPtir i~lclrclcnce from the fan cards (2 AC motors per card) is quite tliffirlllt in the case of a device which shows one count per day background, but electrom~gnrti~ shielding is ~ypruyliate. Cooling elemrntc based on Peltier effect (no AC
~:UllCllki) offer a less ~liffirlllt solution.
Software preferably permits "se~ml~ss" image reconstruction, taking into account the non-active area between detector units, and is consistent with use of coll-l,ul~ls based on the Intel P6 processor.
Multiplexing several SR-PMT based detectors and building a large-area detector requires faster pulse shape analysis. In single detector systems, DSO's with long llal~rer - times (200 microseconds/pulse) are ~eqll~tf~. The large MPD-Imager uses DSO's with a faster conlyulcl interface. To fully utilize this advantage requires fast drivers and accelerated CA 0223629~ 1998-04-29 pulse procescin~. The logic of the pulse shape rejection is thelcrulc opLillli~d for the multiplexed systems by taking into consideration the additional il~lllla~ion from all PMT's and ch~n~ing the order of the tests. Further oplillli~alion of the pulse shape analysis code is achieved by lltili~ing the highly efficient native Pentium and P6 instructions.
Rec~llce the MPD is targeted for qll~ntit~tion/im~ging of very small amounts of radiolabel, st~tictir~l analysis of data from sources with poor signal-to-background ratio (S/B) and cignifir~nt uncertainty is illll oll~llL for correct data hlte.~lctalion. It is thus ~rcr.,llcd that the MPD-Imager inrludes sofhvare ~ lir~l analysis functions and routines for image ellh~-re.~ and recognition. The latter are preferably based on such AI tPchniqlles as sim~ tPd neural llclwulhs and/or ilclalivc use of memory matrices.
Large MPD-Irnager based upon tirne delay techniques: Some limitations of the large MPD-Imager relate to the SR-PMT's used -- high radioactive bac~l~ulld due to c~ ;."~ ion of ~m~m~t~l tubes with K40 and difficulty of calibration due a large dead zone bet~,veen the four Scp~alc SR-PMT's.
Anger c~mPr~c are a popular class of large gamma detectors wherein the surface of a single large NaI(Tl) crystal is populated with many photomultipliers. When a photon is absorbed in the crystal, the light is shared between several PMT's. The center of gravity of the light pulse is established by finding the four PMT's with the largest signal and reconstructing the event from the ratio of their signal amplitudes. Spatial resolution is limited by crystal thirl~nPss, light yield and the number of PMT's used. Typical Anger camera parameters are: NaI(Tl) crystal thirl~nP.cs = 0.5 inch, number of PMT's = 36-64; intrinsic spatial resolution = 2-3 mm. For I'25, the crystal thirknPs.c can be reduced to 1.5 mm, which i~proves spatial resolution down to 1 mm. In the simplest implemPnt~tion~ the number of ADC's is the same as the number of PMT's, say twenty-four 2" PMT's. More complicated srhPm P-s are envisioned according to this aspect of the invention, in which a large number of thresholds are used but only eight ADC ch~nnPlc are nPress~ry. An Anger camera based MPD-Imager is plcfcllcd, but the required ele-;l.olfics are complex and due to the large number of channels, pulse shape analysis is difficult to implement.
An ~ ,.live embodiment is a high pclrollllal~e spatially resolving gamma detector using a pulse delay technique and DSO to reconstruct the photon position, and the new scintill~tor material, yttrium ~ ... perovskite activated with Ce (YAP(Ce)). It provides - - a high light yield (about 50% of NaI(Tl)) and is about five times faster than NaI(Tl). Also, -CA 0223629~ 1998-04-29 WO 97/16746 PCT~US96/16968 mllltirll~nn~l, high speced and large lllClllOly DSO cards comr~tihle with Intel Pentium or P6, e.g., two channel, 150 MHz/channel with 32 kb of memory from Gage Inc., may be used.
Low att~M~3tion analog delay elemPnt~ are readily available with delay times from a few tens of ~noseconds to a few microseconds. With a triangularly shaped YAP pulse (rise time of 10 ns and fall time of 50 ns), the outputs of up to 10 PMT's can be (1igiti7.od with a single channel of DSO. Thus, two DSO cards encode the output of up to 40 PMT's and permit highly reliable, nomin~lly 8 bit q~ntit~tion. One can use modulo four encoding, where the four neighbolillg PMT's are each encoded upon dirr~,.e.ll DSO channels. Thus, a compact, relatively low cost implP~ ion of Anger camera is possible.
Using a YAP scintillator crystal for the MPD-Imager, signal pulses have about a 50 nsec duration. This allows multiplexing signals from several PMT's onto a single DSO input through the use of collsl~l delays. S~ ;llg is plcr~ ,d for the 6 outputs from each of the 4 rows of PMT's delayed by 0, 100, 200, 300, 400 and 500 nsec, lc~peeLi~ely. This pulse train is then sent to a DSO input channel and to the trigger card (SCPC) which produces triggers for all DSO's in the system. For the second (coincidence) detector a NaI (Tl) crystal of the same size (12" x 8") is read out by four 3" PMT's. The outputs of these PMT's are s~mmPd and the sum signal used to establish whether the event is coincident. Additionally, this pulse is pulse shape analyzed to reject background. All thresholds, digital delays, and gains are under software control through the SCPC. The colllplllcl decodes the multiplexed signals acquired by the DSO's and finds the cool~ es of each d~tected event through analysis of the pulse amplitudes in all 24 PMT's.
To avoid the problem of radioactive background, the large MPD-Imager uses carefully selected PMT's based on radioactive background, detection efficiency, energy resolution, homogeneity over the photoç~thod~ surface, dark current, and long term stability. Plercll~,d photom~ltipliers include either 1" or hexagonal 2" PMT's from EMI. For an MPD-Imager of 12 inches x 8 inches, the ~lc~cllcd configuration is a grid of 2" hexagonal PMT's with 2"
center to center spacing, leading to a 6 x 4 array of PMT's. Alternatively, for 1" PMT's spaced 1.5 " center to center, an array of 8 x 5 PMT's is ~lcfcllcd. Cost is much lower for 2" PMT's because the electronics are much simpler and only two DSO cards are needed. On the other hand, the 1 " PMT provides slightly better spatial resolution and about a factor of 2 lower radioactive background.
- The large MPD-Imager preferably uses de~ tPrl data acquisition and proces~ing software. The signals from several PMT's are encoded using delay lines and multiplexed onto CA 0223629~ 1998-04-29 one DSO channel. The acquisition sOflwal~ processes a succe~eeion of pulses using ap~ ia pulse-shape rejection and baseline restoration procedures. Although this task is simplified by the CO~Lall~ delay times belw~ll signals, additional co~ lr~ y analysis of whether the pulses in the "train" are due to a single event, and even some deconvolution may be required. A
reliable and highly efficient process is plefelled for COlllyu~ g the coordilla~s of each event based on the data from multiple PMT's. Special provision is made for storing and processing much larger volumes of data, as well as for co,lll)el~d~ing for the inevitable non-L~nirol."i~
in tbe large-area detector. These louliiles can be O~lhl~i~ed by replacing most colll~uL~Lions by look-up tables, and coding them as much as possible, within the limits of integer ~ ir, 10 in assembly language to ~l;"~;"i~ll the dead time. Pulse processing, ~ecLlulll analysis and image reconstruction are otherwise as for the single-detector SR-MPD system.
Sequential sample MPD device: The ssMPD is a twelve sample, benchtop radiation counter that can be used to measure counts per minute and c~lr~ te ~ einttogrations per minute emitted by multiphoton emission isotopes, including l25I. In volumes of 0.05 to 2.0 mls 15 contained in 12 mm x 75 mm or 13 mm x 100 mm sample tubes dishll~glalions are nleasul~d over the range of 1 to 106 dpms. The sample tubes are capped to avoid cont~min~ting the detector ch~mher.
A layout of the ssMPD is shown in Figure 6. Tube holders 70 are mounted on hori_ontal tube drive 71 lulllling above the lead shield 72. Two detectors 73, each colllpli~ing 20 a scintillator 74, a PMT 75, and base electronics 76, are mounted facing separator 77.
Vertical tube lift 78 lowers and raises each of the sample tubes (not shown) in turn. The detector and mrch~nirs are supported by frame 79 and the entire device is encomraeeecl by enclosure 80.
The detector assembly is a twin detector system coneieting of two scintillator crystals, 25 each coupled to a high resolution planar photomultiplier. The detector assembly is e~r~ePd in a composite lead-tin-copper shield to minimi7P en~dlolllllcbllLal background. In addition to pc..~ li"g efficient counting of gamma ellliLL~l~, the sample holder acts as a separator to reduce crosstalk within the twin scintillator detector assembly.
The detector assembly is connected directly to a personal co~ ulel (PC) configured 30 with ssMPD read-out ele~;Llonics, control and data logger software for WindowsTM, and optionally, a printer. The software outputs, to a printer or a file, a general report header and, for each sample: sample llumbel, sample position, count time, measured counts per minute, c~lrul~ted di~hlLeglaLions per minute, and st~ti~tir~l uncellaill~y. In addition, the software CA 0223629~ 1998-04-29 supports data export to a variety of cu~ uel~;ial data analysis and assay illt~ lc~lion sorlv~al~, p~ gçs.
For l25I, the specific regions of energy and the detection efficiency are established ~ tnm~tir~lly during the calibration procedure. For other isotopes the counting is pelrolllRd S with limited energy and pulse shape disclhl~u~lion (i.e., with a higher background), and the absolute detection errlciell.;y is unknown. If the absolute activity of non-l25I samples is desired, the operator may acquire a calibrated source of the isotope in question and establish the ~letPction efficiency of the ssMPD for this isotope.
The ssMPD system employs twin scintillator crystals with photomultiplier tube (PMT) 10 read-outs as a gamma- and X- ray emissions detector. The read-out electronics and software amplify and shape the signals from each PMT, analyze the pulse shapes and build a combined energy spectrum for subsequ~nt pulse height analysis.
The external ~hiP!~ling of the ssMPD detector ~ s the effects of ambient radiation; therefore, the predominant collll)onelll of the non-radioactive backglou ld is due to 15 dark pulses in the PMTs. The shape of these pulses is dirrelc;lll than those produced by scintillation in the detector, allowing rejection of PMT dark pulses as well as a majority of electrom~gnPtir and vibrational artifacts. After pulse shape based rejection, the backgluulld in the system is almost flat for energies in the 15-100 keV range and is very stable, independent of the activities in the vicinity of the detector (with the exception of hard gamma 20 radiation capable of pel~ll~lillg the ssMPD shielding).
The non-rejected counts in the appro~,lidt~ energy region for the desired isotope are integrated to d~llllille the count rate. Conversion of the count rate (counts per minute or cpm) into the actual activity in the sample (decays per minute or dpm) requires knowledge of the detection efficiency (DE) of the counter for the specific radiolabel.
The DE for each of the two detectors is cl~ P~ sepalalely to improve the ssMPD'scalibration and accuracy; to test system integrity; and to correct for sample holder mi~lignm.ont by coll.~ g the two estim~tPs of the actual activity of the calibration sample.
A direct in(lir~for of the sensilivily (limit of detection) of the ssMPD is its background equivalent activity (BEA)--defined as the activity of a source that produces a count rate equal 30 to the background in the detector. This figure of merit accounts for both the bac~l. und and the detection efficiency of the ssMPD. The background equivalent activity for the ssMPD is 2 picoCuries, or 1 attomole of l25I, while m~int~ining comparable detection efficiency (DE
50%) and .5ignifi~ntly lowering radioactive background (BKG 2cpm).

CA 0223629~ 1998-04-29 The ssMPD's background rejection techniques permit reliable 4~ ion from 1 to 106 cpm (6 logs dynamic range). Above 106 cpm nonlhl~dlily is caused by pulse pile-up in the detector. Although the response is no longer linear in this range, the dead time of the acqllicition system of the ssMPD is non-e~ten~l~ble~ so that counting can be pclr~ cd and the 5 results corrected for pile-up.
Another bellchlop embodiment uses a pick and place robot arm that picks a selected sample from a sample rack, places it in the sample holder, and removes and returns the sample when counting is complete. In both benchtop embodiments, the ch~nging m.och~nic~ is coupled with the data processing equipment to allow for sample counting time to be 0~1i-,-i,.?~l.
In sllmm~ry, it can be seen that a multiphoton detector of the invention provides llUlll~,lOUs advantages. These include low background and high se~ ivi~y, high detection efficiency, high energy resolution, ex~ellent reproducibility and stability, low cost, and small size.
While this description contains many specific details, these should not be construed as 15 limit~tions on the scope of the invention, but rather as examples of plcr~lled embo~limpntc.
Many other variations are possible. The scope of the invention should be dele~ -cl not by the embo~iment~ described and illustrated, but by the claims as broadly construed together with their equivalents.

Claims (54)

We claim:

1. An apparatus for detecting a radioisotope in a sample, comprising:
(a) a detector assembly including two opposed gamma/x-ray detectors for detecting emissions from the radioisotope in the sample when the sample is placed in a sample holder between the detectors, and for converting the emissions into electric output pulses, and (b) a pulse shape analyzer, operatively connected to the detectors, for analyzing the output pulses from each detector and comparing them to predetermined acceptance criteria for the two detectors, rejecting pulses that do not satisfy the predetermined acceptance criteria, and counting output pulses that satisfy the predetermined acceptance criteria, the counting being done separately for coincident and non-coincident pulses.

2. An apparatus according to claim 1, wherein the detector assembly comprises atleast two opposed detectors, at least one of of which comprises a thin inorganic scintillator crystal that produces a signal when impacted with a photon emission from the radioisotope; a separator placed between the scintillators; a shield array isolating the scintillators from external radiation; and a photosensor to amplify the signal from the scintillator.

3. An apparatus according to claim 2, wherein the scintillator is selected from the group consisting of NaI(T1), CsI(T1), CaF2(Eu), and YAP.

4. An apparatus according to claim 2, wherein the scintillators are identical CaF2(Eu) crystals with a thickness less than about 0.5 inch.

5. An apparatus according to claim 2, wherein the photosensor is a photomultiplier tube made of low radioactive material selected to have radioactivity lower than about 10 pCi.

6. An apparatus according to claim 5, wherein the diameter of the photomultiplier is about 2" to about 3", and the diameter of the scintillation crystal is about 7% to about 25%
smaller than the diameter of the photomultiplier tube.

7. An apparatus according to claim 2, wherein a low radioactive background optically transparent window is placed between the scintillator and the photosensor.

8. An apparatus according to claim 7, wherein the window comprises quartz or a high density ( > 4 g/cc ) and high atomic number (greater than about 50) optically transparent material.

9. An apparatus according to claim 7, wherein the window is made of material selected from the group consisting of high-purity GeO2 or germanium-based glass, high density glasses based on lead, PbF2, and undoped bismuth germanite (BGO), and wherein the windows are optically connected to the scintillator and the photosensor with an optical grease that matches the optical properties of the PMT.

10. The apparatus according to claim 5, wherein the scintillator and photomultiplier tube are covered by at least three subsequent layers of nontransparent plastic tape followed by thin metal tape.

11. The apparatus according to claim 10, wherein the metal tapes include a special metal tape with high magnetic permeability, and the assembly is covered by a few layers of copper tape.

12. The apparatus according to claim 1, wherein the detector assembly is placed within a shield of which at least one of the components is a metal having an atomic number greater than about 61, or a compound thereof having a density greater than about 5 g/cc, and the shield materials are selected to have a radioactive background less than about 10 pCi/g.

13. The apparatus according to claim 1, wherein the sample holder is a plastic tray with dimensions of between about 12 x 13 mm and about 75 x 100 mm.

14. An apparatus according to claim 1, wherein both detectors comprise semiconducting detectors with thickness lower than about 5 g/cm2.

15. An apparatus according to claim 1, wherein both detectors are gas detectors,one of which is a spatially resolving detector.

16. An apparatus according to claim 1, wherein the sample holder comprises an encapsulated sample spot and is thin enough and made of low enough density/low enough atomic number material that the sample holder absorbs negligible amounts of X-rays and does not contaminate the detectors.

17. An apparatus according to claim 16, wherein the sample spot is encapsulated with a thin layer of material with very low radiactive background less than about 0.1 pci/cm2 and low atomic number.

18. An apparatus according to claim 17, wherein the encapsulating material is selected from the group consisting of spray, lacquer, acrylic paint, silicon paint, and GE
vernis.

19. An apparatus according to claim 17, wherein the sample is affixed to a solid, flat sheet within an encapsulating pouch made from a thin film selected from the group consisting of berylium films thinner than about 1 mm, plastic films thinner than about 0.5 mm, and aluminum films thinner than about 0.2 mm.

20. An apparatus according to claim 2 for scanning, wherein one of the scintillation detectors is substantially smaller than the other one, and further comprising a thin film of high density/high atomic number heavy metal having a small aperture placed in front of the small detector.

21. A scanning apparatus according to claim 20, optimized for 125I sources, wherein the scintillators are crystals of CaF2(Eu) with thickness less than about 3mm, and diameters between about 0.5 and about 1 inch for the smaller crystal, and between about 2 and about 3 inches for the bigger crystal.

22. A scanning apparatus according to claim 20, wherein the pulse shape analyzerhas two modes, an OR mode accepting both non-coincident and coincident pulses for counting, and an AND mode accepting only coincident pulses for counting; and further comprising a sample mover adapted for holding and moving the sample holder in two dimensions with spatial resolution adequate to present essentially flat samples to the aperture in front of the small scintillation crystal; and a controller that identifies the position of the sample mover and permits correlation with the count rate for each position; and optimizes the time of data acquisition for each sample position such that data acquisition can be stopped when the accumulated data reaches a predetermined signal to background ratio or signal to statistical uncertainty ratio; and the pulse shape analyzer and controller present the acquired data as a two dimensional image.

23. A scanning apparatus according to claim 22, wherein two scans of the samplesare performed, first with low spatial resolution, and second with higher spatial resolution in selected areas in which the count rate is below a predetermined value.

24. A scanning apparatus according to claim 21, wherein the small scintillator is thinner than about 0.1 mm, is sensitive to Auger electrons or very low energy X-rays (E <
10 keV), and has a diameter between about 0.1 and about 0.5 inch.

25. A scanning apparatus according to claim 20, optimized for 125I sources, wherein the small detector is a silicon scintillator or spatially resolving CCD detector thinner than 0.1 mm, is sensitive to Auger electrons or very low energy X-rays (E < 10 keV), and has a diameter between about 0.1 and about 0.5 inch, and the large scintillator is a crystal of CaF2(Eu) with thickness less than about 3mm, and diameter between about 2 and about 3 inches.

26. An apparatus according to claim 1 with spatially resolving ability, wherein at least one of the detectors comprises a thin NaI(T1) scintillator with a high density/heavy metal film affixed to it, the film having a coded aperture pattern, and wherein the scintillator is coupled with a spatially resolving photomultiplier tube having a plurality of outputs each operatively connected to a matched low noise amplifier, the outputs of which are summated electronically into a sum output, and the individual and sum outputs are input to multichannel spectrum analyzers.

27. An apparatus according to claim 26, wherein the amplified output pulses fromthe spatially resolving PMT are compared with "standard shape" pulses according to pulse shape acceptance criteria, and only events satisfying the "standard shape" condition in both the spatially resolved and non-spatially resolved PMTs are accepted and used to build a two dimensional sample image.

28. An apparatus according to claim 6, wherein the spatially resolving photomultiplier has four outputs, and the position of impact of an X-ray is calculated from the iterative algorithm, the first approximation given by x0 = (SL - SR)/TS and y0=(ST - SB)/TS
and following approximations obtained from:
x i = (a[x i-l] * SL - b[x i-l]*SR)/TS
and y i = (c[y i-l] * ST - d[x i-l]*SB)/TS
wherein SL = signal left, SR = signal right, ST = signal top and SB = signal bottom (SB) and TS = SL + SR + ST + SB and where a(x), b(x), c(y) and d(y) are the calibration functions established empirically for the spatially resolving photomultiplier.

29. An apparatus according to claim 28, wherein the true energy of an X-ray is calculated from the impact position of the X-ray using the look up calibration tables established empirically for the spatially resolving photomultiplier.

30. An imaging apparatus according to claim 26, wherein the NaI(T1) crystal has affixed to it a coded aperture mask made of high density/high atomic number film with a regular pattern of up to about 50 holes or slits; the sample holder is a flat two dimensional array of samples; and further comprising a computer-controlled mover which sequentially places samples in measurement positions in front of the aperture mask for a pre-set time.

31. An imaging apparatus according to claim 30, further comprising an imaging analyzer wherein the image is reconstructed from the counting data; the resolution-defining aperture pattern is such that to obtain a continuous image the pixels have to be interleaved in both X and Y dimensions and the pattern allows interleaving; for each available resolution mask the system has a separate calibration file containing the software mask defining the mapping of the apertures to the image surface; the user interface allows selecting a rectangular area to be scanned and pre-setting the counting time for each pixel; and a data presentation system permits visualization of the image in false colors, shades of grey, a contour map or as a 3D surface, and stores the image in a number of standard formats for analysis by other image processing programs.

32. An apparatus according to claim 1, wherein one of the detectors is a Ge- semiconducting detector optimized for detection of X-rays, a silicon detector, a spatially resolving gas detector.

33. An apparatus according to claim 32, wherein the spatially resolving detector is a thin self-limiting streamer chamber, wherein the spatial resolution is obtained by imaging, using a CCD camera.

34. A method for detecting a radioisotope in a sample, comprising:
placing the sample in a sample holder between two opposed detectors;
detecting emissions from the sample using the two opposed detectors to obtain electrical output pulses;
comparing the output pulses to predetermined acceptance criteria for the two detectors;
rejecting output pulses that do not satisfy the predetermined acceptance criteria; and counting output pulses that satisfy the predetermined acceptance criteria, the counting being done separately for pulses corresponding to coincident and non-coincident emissions.

35. An apparatus according to claim 1 wherein:
the detector assembly comprises means for holding samples sandwiched between means for converting the emissions from the isotope to electrical output pulses; and the pulse shape analyzer comprises means for counting and timing the output pulses, means for normalizing output pulse amplitude to fall within a predetermined range over a baseline and to correlate with the energy of the emissions, means for generating a first trigger pulse when there is an output pulse having an amplitude which exceeds a predetermined threshold value, and generating a second trigger pulse when there are two substantially simultaneous output pulses having respective amplitudes which exceed the predetermined threshold value, and rejecting output pulses that do not exceed the predetermined threshold value, means responsive to a trigger pulse for capturing a predetermined number of subsequent normalized output pulses, means for analyzing the captured output pulses based on predetermined isotope and system-dependant acceptance criteria selected from the group consisting of pulse shape, height, coordinates, and coincidence, and means for counting pulses that satisfy the predetermined isotope and system-dependant acceptance criteria and rejecting pulses that do not satisfy the acceptance criteria.

36. An apparatus according to claim 35, further comprising means for shielding the detectors from radioactive background.

37. An apparatus according to claim 35, further comprising means for absorbing the majority of external X-rays.

38. An apparatus according to claim 1, wherein the sample holder has a plurality of conical holes ordered into an easily distinguished pattern, and samples to be studied are placed into the holes.

39. An apparatus according to claim 38, wherein the sample holder aligns samplesblotted upon an appropriate thin film with patterns substantially identical to the hole pattern in the separator with the holes in the sample holder.

40. An apparatus according to claim 39, wherein the sample holder has two parts,each with identical hole patterns, and the blotted film is placed between these two parts.

41. An apparatus according to claim 1, wherein the detector assembly comprises for each detector a preamplifier, amplifier and shaping amplifier.

42. An apparatus according to claim 1 for selective quantitation of a coincidentgamma/X-ray (CGX) emitter in a sample, wherein the detector assembly comprises:
(a) means for detecting coincident gamma and x-ray emissions from the CGX emitter as output pulses in separate radiation detectors, and the pulse shape analyzer comprises (b) means for analyzing the shape and height of the output pulses from the detectors, (c) means for discriminating and rejecting non-coincident output pulses, (d) means for discriminating and rejecting spurious coincident output pulses, and (e) means for using the remaining output pulses to quantify the presence of the CGX
emitter in the sample.

43. An apparatus according to claim 42, capable of detecting less than one attomole of labeled molecule.

44. The method of claim 34, further comprising:
performing a setup of the apparatus by:
determining and storing detector dependent permitted ranges for a plurality of pulse shape rejection parameters for the at least one detector, the parameters being associated with a desired characteristic emission to be detected;
operating the apparatus to detect the desired characteristic emission by:
determining pulse shape parameters of an output signal from the at least one detector; and comparing the pulse shape parameters of an output signal from the at least one detector with the stored detector dependent permitted ranges;
wherein, if the pulse shape parameters of the output signal from the at least one detector do not compare closely with the stored detector dependent permitted ranges, the output signal from the at least one detector is rejected as a detection of the desired characteristic emission.

45. The method according to claim 44, wherein the setup comprises:
providing to the at least one detector a sample of a source of the desired characteristic emission;
acquiring a plurality of pulses from the at least one detector;
building and storing histograms of the plurality of pulses, including current baseline, and pulse widths at 1/4, 1/2 and 3/4 of the pulse amplitude;
determining the detector dependent permitted ranges for a plurality of pulse shape rejection parameters for the at least one detector as those accepting 99%, for each histogram, of the contiguous dominant peak area of the histogram;
storing the determined detector dependent permitted ranges for the plurality of pulse shape rejection parameters for the at least one detector

46. The method according to claim 45, wherein the setup further comprises:
acquiring a plurality of coincident events, a coincident event being substantialcoincidence of output from the first and second detectors;
determining and storing permitted ranges of pulse displacement parameters by building histograms and using the 99% accepting contiguous dominant peak area of the histogram approach; and storing the permitted ranges of pulse displacement parameters.

47. The method according to claim 46, wherein the setup further comprises:
building three separate energy spectra associated with the desired characteristic emission, including a spectrum for the first detector, a spectrum for the second detector, and a two-dimensional spectrum of coincident events.

48. The method according to claim 44, wherein, in the operating of the apparatus to detect the desired characteristic emission, the determining pulse shape parameters of an output signal from the at least one detector comprises:
acquiring a plurality of data points for a received pulse over a time window spanning the whole width of the pulse and a short time prior to the rise of the pulse;
calculating an average of the first few data points to determine a current baseline;
if the calculated current baseline differs significantly from 0, rejecting the pulse as distorted by pile-up;
if the pulse was not rejected in the preceding step, scanning a plurality of next data points to find a maximum value;
if the maximum value found exceeds an overflow amount, rejecting the pulse;
if the pulse was not rejected in any of the preceding steps, calculating a peak amplitude of the pulse as the difference between the maximum value and the current baseline;
scanning forward and backward from the peak of the pulse to determine time offsets for amplitude levels at a plurality of fractions of the peak amplitude; and using at least the determined time offsets as the pulse shape parameters of the output signal from the at least one detector.

49. The method according to claim 48, wherein, in operating the apparatus to detect the desired characteristic emission, the comparing of the pulse shape parameters of the output signal with the stored detector dependent permitted ranges comprises:
determining if the time offsets for amplitude levels of 1/2, 1/4 and 3/4, of thepeak amplitude of the pulse are within the stored detector dependent permitted ranges.

50. The method according to claim 48, wherein the at least one detector comprises a first and a second detector, wherein the setup further comprises:
acquiring a plurality of coincident events, a coincident event being substantialcoincidence of output from the first and second detectors;
determining and storing permitted ranges of pulse displacement parameters by building histograms and using a 99% accepting contiguous dominant peak area of the histogram approach; and storing the permitted ranges of pulse displacement parameters; and wherein operating the apparatus to detect the desired characteristic emission further comprises testing for the occurrence of coincidence during operation, including:
determining the distance between the respective time offsets for the 1/4 amplitude levels on the rising front of the respective pulses from the first and second detectors; and comparing the determined distance with the stored permitted ranges of pulse displacement parameters.

51. The method according to claim 44, further comprising calibrating the apparatus for a particular isotope.

52. An apparatus according to claim 1, wherein the pulse shape analyzer comprises an article of manufacture comprising a computer readable storage medium having a substrate physically configured to represent a computer program, the computer program comprising means for setting-up and operating a programmable particle/radiation emission detection apparatus.

53. An apparatus according to claim 1, wherein the detector assembly comprises:
a scintillator crystal, a photomultiplier optically connected to the scintillator crystal, an integral photomultiplier base electronics module connected to the photomultiplier, comprising a high voltage power supply, voltage divider, and amplifier, shields between the base module and the photomultiplier, between the photomultiplier and the scintillator crystal, around the photomultiplier, and around the scintillator-photomultiplier-base module assembly, all materials used in the detector being of radioactivity below about 1 cpm background.

54. A method for detecting a radioisotope in a sample as in claim 34, further comprising:
normalizing the output pulse amplitude to fall within a predetermined range over a baseline and to correlate with the energy of the emissions;
generating a first trigger pulse when there is a triggering output pulse for which the amplitude of the output pulse exceeds a predetermined threshold value, and generating a second trigger pulse when there are two simultaneous triggering output pulses for which the amplitude of the output pulses exceeds the predetermined threshold value, and rejecting pulses that do not exceed the predetermined threshold value;
responsive to a trigger pulse, capturing a predetermined number of subsequent normalized output pulses; and analyzing the captured output pulses as to predetermined isotope and system-dependant acceptance criteria.